Hematological malignancies (HM) are a group of neoplastic diseases that are usually heterogenous in nature due to the complex underlying genetic aberrations in which collaborating mutations enable cells to evade checkpoints that normally safeguard it against DNA damage and other disruptions of healthy cell growth. Research regarding chromosomal structural rearrangements and alterations, gene mutations, and functionality are currently being carried out to understand the genomics of these abnormalities. It is also becoming more evident that cross talk between the functional changes in transcription and proteins gives the characteristics of the disease although specific mutations may induce unique phenotypes. Functional genomics is vital in this aspect as it measures the complete genetic change in cancerous cells and seeks to integrate the dynamic changes in these networks to elucidate various cancer phenotypes. The advent of CRISPR technology has indeed provided a superfluity of benefits to mankind, as this versatile technology enables DNA editing in the genome. The CRISPR-Cas9 system is a precise genome editing tool, and it has revolutionized methodologies in the field of hematology. Currently, there are various CRISPR systems that are used to perform robust site-specific gene editing to study HM. Furthermore, experimental approaches that are based on CRISPR technology have created promising tools for developing effective hematological therapeutics. Therefore, this review will focus on diverse applications of CRISPR-based gene-editing tools in HM and its potential future trajectory. Collectively, this review will demonstrate the key roles of different CRISPR systems that are being used in HM, and the literature will be a representation of a critical step toward further understanding the biology of HM and the development of potential therapeutic approaches. Hematological malignancies (HM) are cancers that begin in the cells of blood-forming tissues such as the bone marrow or generally in the cells immune system. In the revised World Health Organization (WHO) classification, hematological malignancies are divided according to morphology, immunophenotype, and genetic and clinical features . FurtherThe current advancement in the field of molecular biology has spurred the way toward understanding a disease in an extensive manner and especially deepens our knowledge of hematological malignancies. Many advanced molecular biology and bioinformatics tools such as PCR, NGS, and karyotyping are currently utilized to understand the pathogenesis of hematological malignancies. Nevertheless, these methods only allow us to map genes but not fully understand the function. Therefore, it is imperative for researchers to utilize functional genomics to decipher and unravel new mechanisms that were previously unknown.Advancement in molecular biology has now paved the way for us to understand the roles of genes that are involved directly or indirectly in the molecular and cellular mechanisms underlying hematological malignancies. The way forward now is to take the optimal charge through unraveling the roles of specific genes by manipulating their functions through gene editing. Currently, there are many techniques available to study gene function including site-directed mutagenesis and siRNA-mediated gene silencing .Nevertheless, these techniques are regarded to be less efficient and specific as compared to the various gene editing techniques  which wiGene editing is the ability to make extremely precise changes in the DNA sequence of a living organism and essentially customizing its genetic makeup. Gene editing is achieved by using enzymes, predominantly nucleases, that have been synthesized to target a specific DNA sequence. This is when they introduce cuts into the DNA strands, allowing the removal of existing DNA and the insertion of a replacement DNA. It is a type of genetic engineering tool to insert, delete, or replace a DNA in the genome of an organism using “molecular scissors” . These n2-His2 ZFNs are fusions between a custom-designed Cys2-His2 zinc-finger protein and the cleavage domain of the FokI restriction endonuclease. Cys2-His2 ZFPs bind DNA by inserting an α-helix into the major groove of the double helix (via the zinc-finger DNA-binding domain. One major concern associated with the use of ZFNs for genome editing is off-target mutations (The discovery of zinc-finger nucleases (ZFN) in the 1980s added value to the gene editing approach by being a precision tool in genome editing: it carried a site-specific recognition pattern in editing the desired gene. The ZFNs are composed of two parts, namely, Fokl nuclease that is fused with zinc-finger DNA-binding domains. The zinc-finger DNA-binding domain has a unique characteristic of recognizing a 3-base pair site on DNA, and it can be combined to recognize longer sequences . Furtherle helix  ZFNs funutations . As a reutations . The maiutations . The upputations . TherefoXanthomonas bacteria that naturally alters the transcription of genes in host plant cells. The binding of TALE to DNA is mediated by a central region that contains an array of 33 to 35 amino acid sequence motifs. The amino acid sequence of each repeat is structurally similar, except for two hypervariable amino acids (the repeat variable di-residues or RVDs) at positions 12 and 13. DNA-binding specificity is determined by RVDs, with ND specifically binding to C nucleotides, HN to A or G nucleotides, NH to G nucleotides, and NP to all nucleotides. There is a one-to-one correspondence between RVDs and contiguous nucleotides in the target site, constituting a strikingly simple TALE–DNA recognition cipher.Transcription activator-like effector nuclease (TALEN) has rapidly emerged as an alternative to ZFNs for genome editing, and they are bacterial effector proteins. In 2009, the code used by TALE proteins to recognize DNA was uncovered. TALE DNA–binding domains can be constructed using a variety of methods, with the most straightforward approach being the Golden Gate assembly . TALENs Functional endonuclease FokI is factitiously fused to DNA-binding domains to create site-specific DSBs and thereby stimulate DNA recombination to achieve TALEN-induced targeted genomic modification. To cleave the two strands of the targeted DNA, the FokI cleavage domain must be dimerized. Hence, like zinc fingers, such a TALEN module is designed in pairs to bind opposing DNA target loci, with proper spacing (12–30 bp) between the two binding sites. However, compared to zinc-finger proteins, there is no need to redesign the linkage between repeats constituting long arrays of TALEs, whose function is to target individual genomic sites. Following pioneering studies on zinc-finger proteins, multiple effector domains have become accessible to support the fusion of TALE repeats for different genomic modification purposes, including nucleases, transcriptional activators, and site-specific recombinases. Although their simpler cipher codes provide better simplicity in design than triplet-confined zinc-finger proteins, one of the primary technical hurdles for cloning repeat TALE arrays is the design of identical repeat sequences on a large scale. To address this limitation, a few strategies have been established to facilitate the fast assembly of custom TALE arrays, including “Golden Gate” molecular cloning, high-throughput solid phase assembly, and connection-independent cloning techniques. More recent advances in TALEN assembly, though, have focused on the development of methods that can enhance their performance, including specificity profiling to uncover non-conventional RVDs that improve TALEN activity, directed evolution as means to refine TALE specificity, and even fusing TALE domains to homing endonuclease variants to generate chimeric nucleases with extended targeting specificity . TALENs cas gene content, CRISPR-Cas systems can be classified into six major types (I–VI)  technology represents a significant improvement over these, reaching a new level of targeting, efficiency, and ease of use. CRISPR nucleases lead the gene-editing platform because they are the most powerful and direct to use gene-editing tools available now. Based on their s (I–VI) . The syss (I–VI) . The spes (I–VI) . It alsoS. pyogenes Cas9, and it still remains to be the most popular CRISPR nuclease for the gene editing approach. The SpCas9 is a colossal CRISPR nuclease bearing 1,368 amino acids in size and possesses a moderately relaxed protospacer adjacent motif  as compared to its orthologs. SpCas9 happens to be the most well-characterized of all CRISPR systems due to it being the first to be repurposed as a gene editor. Thus, making it readily available in virtually any format  and in-line with other genome engineering applications. In bacteria, the type-II CRISPR system provides protection against DNA from invading viruses and plasmids via RNA-guided DNA cleavage by Cas proteins (trans-activating crRNA (tracrRNA) to direct sequence-specific degradation of pathogenic DNA by the Cas9 protein. This system has since been simplified for genome engineering and now consists of only the Cas9 nuclease and a single guide RNA (gRNA) that contains the essential crRNA and tracrRNA elements  and C. jejuni (CjCas9), both of which can be packaged in AAV. SpCas9-NG is engineered with valine to disrupt the interaction between Cas9 and the second G of the 5′-NGG PAM. There is additional modification done to this SpCas9-NG based on molecular modeling to restore the cleavage activity of this “relaxed” Cas9 variant , SpCas9 nickase (Cas9n), and nuclease-dead SpCas9 (dCas9). In order to differentiate these ortologs from SpCas9, there are two main characteristics which are size and PAM specificity. The size really matters for hard-to-transfect cell lines which are vectors that easily exceed 10 kb as well as AAV applications where AAV can only package ∼4.5 kb. Compact Cas9 ortologs include those isolated from  variant . The sec variant . The dCa variant .Streptococcus thermophilus). Relaxed PAMs are good for more flexibility in the gRNA design, but it should be avoided because it may lead to off-target activity. When the condition allows for increased gRNA-binding sites, it will lead to higher potential off-target activity. However, like any emerging technology, the CRISPR-based assay system has minor potential pitfalls, including promiscuous off-target activity by Cas9. To address this issue, various preventative strategies have been employed, such as introducing purified Cas9 straight into target cells, using Cas9 nickase (Cas9n), decreasing sgRNA sequences by 2–3 nt, and exploiting additional two guanines at the 5ˊ terminus of gRNA immediately juxtaposed to the target-complementary region  binding sites there are in a given genome for a given PAM. Ortologs with simple PAMs can interact with more genomics sites compared to those with stringent PAMs. For example, there are more instances of 5′-NGG in any given genome as opposed to 5′-NGGNG which is the PAM site for St3Cas9  domains that target single-stranded RNA (ssRNA). It encompasses a functionally distinct nuclease that is responsible for catalyzing crRNA maturation to form a Cas13a:crRNA complex that is competent for target RNA binding. The binding of the said complex to a complementary ssRNA is termed as activator-RNA. This binding activates the HEPN-nuclease for both target and general ssRNase activity. The action of the HEPN-nuclease is repressed by fractional occlusion of the HEPN active site until binding to the activator-RNA occurs. This effectively makes the activator-RNA the allosteric switch for RNase activity (via RPA or reverse transcriptase RPA (rt-RPA), before incubating the sample with Cas13a and reporter probes, and fluorescence is then measured. SHERLOCK can detect nucleic acids in patient biofluids down to low atto molar concentrations, allowing the detection of tumor mutation in cell-free DNA (cfDNA). In mock cfDNA samples, SHERLOCK can detect two cancer mutations, EGFR L858R and BRAF V600E, under low allelic fraction with single-base mismatch sensitivity. In addition to in vitro RNA target detection, catalytically inactive Cas13a retains its RNA-binding activity such that it can be coupled to a fluorescent probe to enable live cell RNA tracking. This provides an alternative method to recognize and visualize RNA  Cas9 combined with engineered single guide RNA (sgRNA) scaffolds that bind to fluorescent proteins. In this applied science technique, it allows simultaneous imaging of up to seven chromosomal loci in an individual human cell and observes large differences in the chromodynamic properties of different chromosomal loci at a time. This system is a valuable tool for studying the transfiguration of the genome in real time basis, and it is used to label the DNA and then track the movement of DNA in live cells .In summary, the CRISPR system allows for site-specific genomic targeting in virtually any organism . It alsoAML is a multifaceted genetic disease caused by an interwoven process in which numerous collaborating mutations that allow a cell to evade the checkpoints that normally safeguard it against DNA damage and other disruptions of healthy cell growth . It is ain vitro study of both the phenotypical cellular changes induced by such FLT3 mutations and the different levels of response to anti-AML drugs, including the potential for discovering new therapeutic targets for AML treatments. A number of researchers have utilized CRISPR systems to investigate the genetic targets behind differentiation blockages commonly seen in causing the development of AML. For instance, Wang et al., identified RNA-binding protein ZFP36L2 as a crucial mechanistic regulator of cell differentiation in AML using cell surface antigen–guided-CRISPR systems  is a malignant, cancerous development in hematopoietic stem cell lines attributed by several gene mutations, CRISPR serves as an excellent tool to study the underlying molecular and genetic reasons for AML progression. Researchers such as  utilized systems . Geneticchanisms .et al., applied the gene-editing process of CRISPR/Cas systems in knocking out interleukin-1 receptor accessory protein (IL1RAP) genes in human leukemia stem cells in an attempt to abolish relapses of leukemia in AML patients by designing a bone marrow scaffold–mediated delivery of the therapeutic CRISPR system  therapies for effective patient treatments , thus increasing the chances of eliminating this cancer in patients. A latest study conducted by Tzu-Chieh Ho et al., showcased scaffold-mediated CRISPR-Cas9 delivery system for targeting the gene interleukin-1 receptor accessory protein (IL1RAP) in human LSCs . This apin vitro and in vivo. Altogether, the data demonstrated the utility of CRISPR/Cas9 screens to uncover mechanisms underlying the sensitivity to DNT therapy and suggests CD64 as a predictive marker for response in AML patients (Acute myeloid leukemia (AML) remains a devastating disease in need of new therapies to improve patient survival. Targeted adoptive T-cell therapies have achieved impressive clinical outcomes in some B-cell leukemia and lymphomas but not in AML. Double-negative T cells (DNTs) effectively kill blast cells from the majority of AML patients and are now being tested in clinical trials. However, AML blasts obtained from ∼30% of patients show resistance to DNT-mediated cytotoxicity; the markers or mechanisms underlying this resistance have not been elucidated. Here, a targeted clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) screen was utilized to identify genes that caused susceptibility of AML cells to DNT therapy. Inactivation of the Spt-Ada-Gcn5-acetyltransferase (SAGA) deubiquitinating complex components sensitized AML cells to DNT-mediated cytotoxicity. In contrast, CD64 inactivation resulted in resistance to DNT-mediated cytotoxicity. Importantly, the level of CD64 expression correlated strongly with the sensitivity of AML cells to DNT treatment. Furthermore, the ectopic expression of CD64 overcame AML resistance to DNTs patients .ALL can be characterized by looking at the development of precursor lymphoid into B or T cell. This type of leukemia is usually rare but requires immediate treatment as ALL leukemogenesis is rapid and aggressive . AccumulThe CRISPR/Cas9 knockout system was utilized in the manipulation of ALL’s genomics and in studying the differentiation of ALL into erythrocytes. A study conducted by Xie and group reported that GATA-1 motif flanking 5 kb upstream ARHGEF12 was impaired due to the CC to TT SNPs in hematopoietic cells of ALL patients suggesting that GATA-1 transcriptional activation of ARHGEF12 is crucial in driving erythrogenesis in which ALL patients lacked. The luciferase reporter assay confirmed GATA-1 interaction onto the ARHGEF12 flanking region when compared to the impaired motif brought about by the CC to TT SNPs. To further ascertain that this switch was responsible in driving erythropoiesis, the group employed CRISPR/Cas9 knockout where guide RNA was designed to be specifically targeting the ARHGEF12 gene directly. Result showed a stunted production of red blood cells and immature erythrocytes accumulation .via the use of CRISPR system was also demonstrated when the MYB-binding site located proximally upstream of LMO2 was targeted using Cas9. In this study, a second binding motif of MYB was found to occur on LMO proximal site, this occurs due to the translocation event of the LMO2 gene which shift the MYB-binding site to a second MYB-binding site thus duplicating the MYB motif resulting in an increase in LMO2 activation by MYB. Interesting enough, Cas9 cleavage of the duplicated MYB motif resulted in a significant reduction in LMO2 expression  treatment, suggesting that ARID5B could probably be downregulated in ALL for chemoresistance. This was interesting as patients with relapse showed significant downregulation of ARID5B . This stAdditionally, ALL metabolism dysregulation was also studied in further understanding the mechanism of ALL leukemogenesis, and it was reported that the PAX5 transcription activator was deactivated in which this trailed increase in glucose uptake of ALL thus supporting ALL development. Using the CRISPR activation system (VP64), the same group induced PAX5 expression which in turn would activate PAX5 repressed gene; this attenuated glucose uptake thus restoring the gate keeping function of PAX5 as a metabolic switch .This type of leukemia can be characterized by looking at accumulation of granulocyte within the bone marrow. Trailing myeloid precursor aberration neutrophilic and basophilic cell accumulation could also be observed within the peripheral blood. Synonymous to CML is the BCR-ABL fusion oncoprotein resulting from the Philadelphia chromosome, which is a direct result of translocation between chromosome 9 and 22 .The use of CRISPR in CML is mainly on aberating the BCR-ABL fusion tyrosine kinase. BCR-ABL fusion resulted in a surface protein with constitutive tyrosine kinase activity which is one of the main factors in CML leukemogenesis . AlthougAn interesting study sought to abolish the region of fusion by specifically targeting the ABL exon 6. In this case, the CRISPR knockout system was used where sgRNA were designed to nick a region of about 100 bp on ABL locus within the genome. The CRISPR nickase system was proven to be effective as expression of the BCR-ABL mRNA reduced to only 2.6%, with a total abolishment of protein translation. This also leads to an increase in apoptosis when compared to wild type CML cells. Furthermore, the group also targeted leukemic stem cells (LSCs) using the same system and found out that mice with LSCs-CRISPR–containing engraftment showed a reduction in CML oncogenic properties and restoration of multipotency of LSC with unbiased development toward only myeloid lineage .E. coli exonuclease 3; this resulted in an efficient cleavage of large genomic locus which is more efficient than CRISPR nickase. In CML, this technology was used to target the BCR-ABL gene where an efficient deletion of the target site was observed, and this was also followed by an increase in apoptosis in CRISPR-EXO3–targeted CML cell line  with either TP53 or ATM mutation. Using CRISPR/Cas9, the group induced deletion of the 11q arm and designed small guide RNA against TP53 and ATM. Results showed that del (11q) with TP53 mutation confer leukemic survivability suggesting that these two mutational events co-occurrence potentiate CLL progression. However biallelic mutation of ATM and TP53 result in reduction in CLL engraftment with CLL . Targetiwith CLL .CRISPR has also been utilized to study the functional genomics of other types of hematological malignancies other than AML, ALL, CML, and CLL. Other common hematological malignancies are multiple myeloma, Hodgkin, and non-Hodgkin lymphomas. Researchers usually conduct whole genome sequencing (WGS) to identify crucial genes. Next, they utilized CRISPR to screen the genes identified with WGS to uncover the function of the genes and how it contributes to the malignancies. CRISPR is also used to uncover how the malignancies adapt and develop resistance to cancer treatments. It is hoped that by studying the functional genomics, new treatment targets against these malignancies can be identified and new treatment options can be developed, tabulated in DLBCL is the most common non-Hodgkin lymphoma worldwide and is a fast growing cancer which affects the B lymphocytes . DLBCL hin vivo researches utilizing CRISPR to elucidate the function of genes in B-cell lymphomas. Felce et al. conducted research on murine A20 lymphoma cell line which is B-cell lymphoma derived from mice as a model for B-cell lymphoma. The researchers utilized CRISPR to knockdown the Foxp1 expression in the cells which upregulated cell surface I-Ab (MHC-II) expression without impairing cell viability in vitro. The reduction of Foxp1 restores immune surveillance. This shows that Foxp1 helps the lymphoma cells evade the immune system. Based on the results, it is hoped that targeting Foxp1 in B-cell lymphomas will enhance the effects of other immunotherapies. By utilizing CRISPR in tandem with other molecular techniques, researchers were able to detect and determine the function of genes crucial in DLBCL  . There ain DLBCL .in vitro and in vivo study uncovered the role of ID3 in deregulation of TCF3 and TCF4 which affects the cell proliferation in BL cells . The results of this study shows that both Sp3 and Phip behave as tumor suppressors in Eμ-Myc driven lymphomas (in vitro and in vivo from BL genome sequencing data.Burkitt’s lymphoma (BL) is another type of non-Hodgkin lymphoma. It is divided into three subtypes which are sporadic, endemic, and immunodeficiency associated . Panea eBL cells . Katigbaymphomas . Both thMultiple myeloma (MM) is the cancer of the plasma cells and is relatively rare cancer that contributes around 1.8% of all diagnosed cancer in United States . There avia loss of surface antigen expression. In the study, CRISPR interference and CRISPR activation were used to identify genes that led to increase in BCMA on the MM cell surface. They discovered that the knockdown of HDAC7 and SEC61A (part of SEC61 complex) increased the levels of BCMA  which have poor clinical outcomes. The goal of the study was to identify targetable vulnerabilities in TCL using genome-wide CRISPR screening. They discovered that potential treatment targets JAK2 or IKZF1 which can be targeted with available inhibitors . In TP53BBC3 (PUMA) is responsible for the resistance of NUTLIN3A in lymphomas. Furthermore, CRISPR-Cas9–mediated silencing of the BBC3 gene confirmed that BBC3 expression might predict NUTLIN3A treatment outcomes in patients. More recently, a study reported the application of a CRISPR-Cas9 system to disrupt CXCR4 expression in mantle cell lymphomas (MCL), a highly aggressive subset of B-cell non-Hodgkin lymphomas (NHLs). Performing lentiviral-based CRISPR-Cas9–mediated silencing of CXCR4, the researchers found that reactive oxygen–mediated CXCR4 expression is a key signal inducing autophagy, which contributes to the survival of bortezomib-resistant MCL cells  cells . This isnografts . This wovia complete knockout  and LexA DNA-binding protein to generate LexA-Fkh1p, which is capable of efficiently recruiting exogenous DNA to the cleavage site  to introduce foreign GOIs into the human genome, low efficiency has been a major obstacle. Knocking-in genes with CRISPR/Cas9 is able to circumvent the detrimental effects on cellular phenotype caused by random integration with lentiviral systems. This approach has many potential uses including the possibility of creating disease models by incorporating mutant genes and also determining the functions of promoters and repressors in knock-in studies. Studies aimed at increasing HDR events have investigated approaches such as using blocking mutations through the incorporation of silent mutations in the PAM and target gene sequence to prevent re-cutting by the Cas9 nuclease once the donor DNA has been ligated . Anotherage site . Furtherage site . This deCurrently, researchers are also highly reliant on cell lines to conduct pooled screenings. Doubts have been cast on the ability of cell line studies to correlation with actual human diseases especially involving functional genomics as cultures maintained for prolonged durations accumulate point mutations, acquire epigenetic modifications, and also develop chromosomal aberrations . The ideAdequately addressing the abovementioned issues will greatly enhance the utility of CRISPR and broaden applications to other emerging areas. Moreover, confounding results occurring from off-targets, false positives, false negatives, and failure of phenotype development could be further prevented to make CRISPR a more robust system in the interrogation of functional genomics. Additionally, it is to be noted that regardless of the hematological malignancies, the potential applications of the CRISPR on gene expression studies remain the same.Advancements in CRISPR/Cas9 technology have greatly impacted basic research in hematological malignancies which have immense potential to be translated into clinical applications in the near future. Among the most promising are in the areas of drug discovery, identification of treatment resistance, disease modeling, and genetic manipulation to improve other cancer therapies such as CAR-T cells and editing HSCs for autologous hematopoietic reconstitution.PAX5 and IKZF1 are among the most commonly altered TFs in B-ALL, affecting approximately 80% of patients  . Furtherpatients . CRISPR hibitors . This aphibitors .via the intrinsic pathway. The information generated from such screens could potentially be used to design combinatorial therapies to simultaneously target the resistance and vulnerable survival genes, thereby reestablishing treatment susceptibility.In identification of drug resistance genes, a genome-scale CRISPR KO screen has been explored in BCR-ABL1 CML cell lines . This apin vivo. In these terms, gene editing with the CRISPR system has presented a simple and versatile solution to leukemic disease modeling with the capability of introducing multiple deletions and insertions to simultaneously manipulate expression of several genes and alleles  T cells. At present, CAR-T cells have been investigated to treat ALL, CLL, and non-Hodgkin lymphoma through the ectopic expression of CD22, CD20, CD19, and dual B targeting . Howeverex vivo HSC modifications. Additionally, this system is also able to couple with conventional gene therapy to guide donor DNA to safe harboring sites for HDR gene integration. These possibilities open new avenues for researchers to explore not only just silencing and activation of expression but also gene replacement as potential therapies for leukemia. As applications of the former methods are obvious, the latter approach will be highly valued in replacing essential genes involved in complex translocations which gave rise to the hematological malignancy. Early studies on correcting the ASXL1 mutation in the CML cell line KBM5 has already confirmed the restoration of gene function and prolonger survival of xenograft mouse models  and PAM sites adjacent to target loci. If there are more than three mismatches between target sequences and 20 nucleotides of gRNA, it can result in off-target effects. Researchers have proposed two types of off-target effects, the first types of off-target effects likely to occur due to the sequence homology of the target loci and the next types of off-target sites occur in the genome other than the target site . The effThe molecular scissors known as “CRISPR” provides complete silencing, and knockdown of gene can still be a valuable means depending on the type of malignancy. This method of gene silencing usually involves an extended time frame to generate a stable cell line. Taking into consideration on the other gene editing tools available with the likes of ZFN, TALENs, and meganucleases, it can be said that CRISPR is the most efficient, stable, and effective. CRISPR is appropriate for delineating gene functions, and genome-editing technology is the apparent option for creating a true genetic null allele, introducing a point mutation, and correcting a specific mutation.In conclusion, through current advancement in terms of gene editing, we can say that CRISPR has taken the research world by storm, evidently allowing us to make changes to the once impossible. CRISPR has made its way onto the freezers of labs all around the spectrum, easily accessible for researchers to use it. It is also becoming a mainstream methodology to study cancer biology given its versatility. It has now matured its way from experimental approach toward customized treatment involving cancer patients. CRISPR gene editing tools have also sparked significant advancements in enhancing our knowledge regarding hematological malignancies, which in essence presents us with potential therapeutic applications which holds much promise for alternative treatments for patients with this group of malignancies. Development of CRISPR utilization in studying the functional genomics of haematologic malignancies is depicted in Figure 1 and a general summary for studying functional genomics via CRISPR based tools is presented in Figure 2.


Anti-glomerular basement membrane disease (Anti-GBM), previously known as Goodpasture syndrome, is an extremely rare cause of rapidly progressive glomerulonephritis and chronic kidney disease stage 5 (CKD5) in children. It is associated with acute pulmonary haemorrhage and it has a poor prognosis. It is classified as an autoimmune, small-vessel vasculitis caused by autoantibody formation against the alpha-3 chain in type IV collagen found in the glomerular basement membrane. Evidence of anti-GBM antibodies in serum or histologically are required for diagnosis. Treatment in children is based on very limited adult data and often involves the use of acute apheresis to rapidly remove circulating factors coupled with intensive immunosuppression such as cyclophosphamide and intravenous corticosteroids. There is also an emerging role for the use of biologic agents such as B cell depletion. The evidence base in children with anti-GBM disease is extremely limited. Multi-centre international collaboration is required to provide insight into this disease, better describe its prognosis and work towards improving outcomes. This review article summarises the key features of this disease in children, highlights treatment options and considers areas of unmet need. Anti-glomerular basement membrane disease (Anti-GBM), previously known as Goodpasture’s disease or syndrome, is an extremely rare cause of glomerulonephritis (GN) and chronic kidney disease stage 5 (CKD 5) in children. The literature relating to anti-GBM disease in children is limited to a small number of case reports and retrospective case series. Although rare, it is characterised by rapidly progressive glomerulonephritis (RPGN) and it is associated with a poor prognosis . This cord and 6th decades of life [Anti-GBM disease is extremely uncommon in children; however, it is responsible for ~ 20% of all causes of RPGN. Defining its precise incidence in children remains a challenge. In adults it has an incidence of 0.5–1.0 cases per million population per year [ of life . The US  of life . Other s of life . The spa of life .The disease classically presents with rapidly progressive GN in 80–90% of cases necessitating acute kidney replacement therapy and this presentation is similar between adults and children with the disease . Most paDRB1*1051 and DRB1*1502 while HLA-DR7 and DR1 appear to be somewhat protective [Detection of anti-GBM antibodies, either in serum or histologically, assist in formulating the diagnosis . In approtective .The glomerular basement membrane is a critical part of the glomerular filtration barrier. Type IV collagen is a major contributor to maintaining the stability of the GBM . Anti-GBANCA-associated vasculitis (AAV) and anti-GBM are included in the spectrum of pulmonary-renal syndromes as they may present with a similar combination of kidney and lung manifestations . Cases oIn patients with Alport syndrome, genetic mutations coding for the type IV collagen chains are responsible for progressive kidney disease . Cases oThe management of anti-GBM disease in children is based on the approach derived from the management in adult disease and clearly described within the recent KDIGO glomerular disease guidelines (2021) . PrincipAnti-GBM is a recognised indication for the use of apheresis treatment in cases presenting with acute GN not requiring dialysis who have diffuse alveolar haemorrhage, or dialysis dependent acute GN without diffuse alveolar haemorrhage . Even inFirst-line immunosuppressive therapy in this disease is cyclophosphamide and high dose corticosteroids as recommended by the KDIGO guidelines for the management of GN . Cyclophn = 119), a third of patients required intensive care unit admission on first presentation, 78% needed kidney replacement therapy (KRT) and 82% received plasma exchange therapy. The 1-year survival rate was 95% and the 3-year survival rate was 92% [Unlike many other autoimmune conditions, anti-GBM disease does not tend to run a relapsing, remitting course. The initial presentation is therefore usually responsible for disease-associated morbidity and mortality. In a large series of adult patients ( was 92% . In this was 92% . The lit was 92% . Graft l was 92% , 40.The evidence base for the diagnosis and management of anti-GBM disease in children is derived from adult data. The proposed standard treatments are based on adult studies and seem to be well accepted . FurtherAnti-GBM disease is a rare entity in childhood but it is associated with a significant level of morbidity and acute mortality. The autoimmune development of antibodies towards type IV collagen in the glomerular and alveolar basement membranes leads to patients typically presenting with RPGN and pulmonary haemorrhage. Evidence of anti-GBM antibodies in serum or histologically is required for diagnosis. Treatment in children is based on adult data and involves the use of apheresis and intensive, strong immunosuppression such as cyclophosphamide. Multi-centre international collaboration is required to provide insight into this disease and to improve its management in children.


Various pathogens have been associated with initiation or exacerbation of neurological diseases such as Alzheimer’s disease and multiple sclerosis. This Research Topic collects knowledge acquired with animal and human studies about the complex host-pathogen interactions during neurological, cognitive, and psychiatric disorders.Zhang et al. focuses on NLR family caspase recruitment domain containing 5 (NLRC5), which has been shown to contribute to a wide variety of immune responses and has recently been connected to neuronal development and central nervous system (CNS) diseases. Here, the authors summarize recent progress on the structure, expression, and biology of NLRC5, its contribution to CNS function and involvement in neurological diseases.Pathogens use unique molecular strategies altering the neuroimmune crosstalk, which initiate from the periphery or directly act in the central nervous system (CNS). Nucleotide-binding oligomerization domain-like receptors (NLRs) are a family of conserved cytosolic receptors that recognize both host- and pathogen-derived intracellular danger signals and play an important role in shaping host defenses and inflammation . The revDow presents evidence supporting the hypothesis that Mycobacterium avium susp. paratuberculosis contributes to dysfunctional autophagy in Alzheimer-infected patients, which in turn may result in neurodegeneration or neurodegeneration-like symptoms. The fact that anti-mycobacterial agents such as rifampin appear to have a neuroprotective role not only in Alzheimer’s but also in Parkinson’s disease  in conferring risk for developing multiple sclerosis (MS) in Sardinia, a Mediterranean island characterized by peculiar genetic polymorphisms and high MS prevalence. Research evidence from studies with humans and animal models show that pathogens may manipulate gene expression in susceptible hosts, leading to immune dysregulation and tissue damage Some studies suggest a link between infections and CNS disorders. In a review,  disease , supporte damage . Molecule damage . MoreoveDuarte et al. leverage the widely used experimental autoimmune encephalomyelitis (EAE) model to study MS and evaluate the effect of asymptomatic brain infection by HSV-1 on the precipitation and severity of EAE in mice. Using both WT and mutant HSV-1, the authors find that HSV-1 infection is associated with more severe EAE in mice compared to uninfected control mice. Their findings support the idea that HSV-1 infection can accelerate and/or exacerbate EAE, which suggests a potential contribution of asymptomatic viral infection to the onset and severity of MS.Although progress has been made in understanding the underlying biology of MS, almost nothing is known about how this disease is triggered. It has been proposed that viral infections including herpes simplex type I (HSV-1) may contribute to precipitation and/or enhancement of MS . Duarte Bellucci et al.). Here, Bellucci et al. take a close look over an 18-month period of SARS-CoV-2 pandemic through the lens of MS and discuss the neuroinflammatory and demyelinating mechanisms associated with COVID-19, review pathophysiological crosstalk between MS and SARS-CoV-2 infection, and investigate SARS-CoV-2 vaccination in the context of MS.Recently, the relationship between chronic viral infection and MS has been complicated by the novel coronavirus, SARS-CoV-2, as its severity of infection heavily depends on host response to infection mirroring several aspects of MS pathobiology  have been tested . The evidence that certain microbes may lead to the expansion of Treg frequency opens to the possibility of therapeutic interventions reprogramming immune balance with pathogens or their derived products.Neuroinflammation is a complex response to CNS injury, which involves the interaction between elements of innate and adaptive immunity. Among adaptive immune cells the regulatory T cells (Tregs) are a major determinant of immune tolerance. Parodi and Kerlero de Rosbo). Altered microbiota and increased intestinal permeability have been associated with pathogenesis of MS and its model, experimental autoimmune encephalomyelitis (EAE) (Parodi and Kerlero de Rosbo sort through the evidence supporting two possible consequences of the gut-brain axis dysfunction in MS and EAE: a) a proinflammatory pro-inflammatory environment and “leaky” gut induced by dysbiosis contribute directly to MS Neuroinflammation; b) neuroinflammation precipitates intestinal inflammation as disease progresses.The gut-brain axis clearly plays a crucial role in neurological diseases, including MS (is (EAE) . HoweverFoster et al. summarizes the evidences about changes in gut microbiome associated with psychiatric conditions and underlines how major depressive disorders present with changes in several immune markers indicating a clear peripheral pro-inflammatory setting under this condition. Moreover, several studies indicate that exposure to various types of stress may alter immunity and gut microbiome and that, vice versa, the microbiome can regulate the impact of stress . Cinthia Farina's research is supported by Italian Ministry for Health (Ricerca Corrente and RF-2018-12367731), FISM  and cofinanced with the 5 per mille public funding.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Furthermore, the development of novel therapeutics for cancer immunotherapy, autoimmune- and inflammatory diseases is tightly restricted by the use of human samples before moving to clinical trials, which is generally slow and costly. Human Immune System (HIS) mice - immunodeficient mice reconstituted with a human immune system – offer the unique opportunity to comprehensively study human hematopoiesis, infectious diseases, autoimmunity, and anti-tumor immunity in vivo. Recent progress in HIS mouse models, in which recipient immunodeficient mice carry several gene mutations or express human cytokines and self-recognition molecules, have improved human hematopoietic stem and progenitor cell (HSPC) engraftment as well as functional immune cell development in primary and secondary lymphoid tissues.Gaining knowledge of human physiology and pathophysiology is often hampered by restricted access to human tissues or limited to performing The articles in this Research Topic describe the latest developments in the field and indicate future directions to further improve HIS models and to apply them for more precisely characterizing human hematopoiesis as well as the human immune response against cancer.in vitro high-throughput screens or gain/loss of function library studies in order to identify novel therapeutic targets for human diseases. The advantage of in vivo models is the more faithful recapitulation of human disease. Xenograft models support the development of human disease, the assessment of patient heterogeneity, and the investigation of the human immune system in vivo. They also allow the preclinical testing of novel therapeutic approaches.Human cell lines and organoids have proven to be particularly useful for Martinov et al. provides up-to-date information on current mouse models that have a humanized hemato-lymphoid system. The authors discuss the underlying principles of these models and point out outstanding challenges to build the next generation of mice with a human hematopoietic and immune system. Importantly, this review also provides guidance on how to identify humanized mouse models suited for specific research investigations. In another review, Mian et al. focus on humanized mouse models to investigate cancer immunotherapy by first giving an overview of recently developed models. In particular, they discuss challenges related to the development of specific human immune cell types and link the assessment of cancer immunotherapies with specific humanized mouse models for preclinical in vivo testing.In the presented series of articles, we aimed at covering the most recent development in the field of HIS mouse models with a particular focus on the application of immunotherapies. The review by Kit gene allow the engraftment of human HSPCs without irradiation due to the reduced competition of human HSPCs with mouse HSPCs for niche space. Hess et al. investigated the engraftment potential of various hematopoietic stem cell sources in non-irradiated NOD.B6 scid Il2rg-/-KitW41/W41 (NBSGW) mice, which carry a hypomorphic mutation of the KIT gene. They show that the human engraftment level in mice transplanted with mobilized adult peripheral blood stem cells is significantly higher than in mice transplanted with human cord blood or bone marrow. The different HSPC sources also yielded different hematopoietic lineage engraftment patterns. However, the transplanted human cells lost the long-term engraftment potential in vivo. Adigbli et al. also used non-irradiated NBSGW mice, demonstrating engraftment of multiple human lymphoid and myeloid cell lineages after transplantation with CD133+ HSPCs. Moreover, NBSGW mice supported the development of human red blood cells when endogenous mouse macrophages were removed. Combined, the two studies provide useful information on the utility of the NBSGW mouse model for studying human hematopoiesis in vivo.Hematopoiesis occurs primarily in the bone marrow, where the hematopoietic stem and progenitor cell (HSPC) undergo expansion and multilineage differentiation. This process is regulated by a specific microenvironment called the hematopoietic stem cell niche. To establish HIS mice, opening the niche by, e.g., sub-lethal irradiation, is required for the stable engraftment of human HSPCs in the physical space. Mice engineered to carry a mutation in the Marín-Jiménez et al., the authors provide a very detailed description of their extensive experience with testing immunotherapies in human cord blood-derived Balb/c Rag2-/-Il2rg-/-SirpaNOD mice. They investigated the response to ICI in various human solid tumors. Their work demonstrates that patient-derived xenografts are suited to investigate immunotherapies based on T-cell responses and constitutes an important resource for the development of preclinical models. Along these lines, Wunderlich et al. developed a preclinical model for pediatric and adolescents and young adults refractory B lymphoblastic leukemia (B-ALL) using NOD scid Il2rg-/- (NSG)-hSCF/IL3/GM-CSFTg or NOD Rag1-/-Il2rg-/- (NRG)-hSCF/IL3/GM-CSFTg mice. Mice were “pre-engrafted” with human cord blood in order to generate functional effector T-cells. They confirmed the clinical efficacy of blinatumomab and also showed that interestingly the application of pembrolizumab reduced the B-ALL burden. Importantly, mice which were not pre-engrafted with human T-cells were unresponsive to treatment. Finally, they observed a synergistic effect of the combined treatment with blinatumomab and pembrolizumab. The combination led to an increased number of mice which lacked minimal residual disease and to prolonged survival supporting further investigation of this dual immunotherapeutic approach. Similarly, Qiao et al. developed a human tumor cell-line derived xenograft model for non-small cell lung cancer in order to test ICI. For this purpose, human PBMCs were transplanted before the tumor injection. They could show that the lactate dehydrogenase inhibitor oxamate and pembrolizumab act synergistically against tumor cells by increasing the number of CD8+ T-cells that infiltrate the tumor microenvironment.Immune checkpoint inhibitors (ICI) are widely used in the treatment of solid tumors. However, a preclinical model to evaluate the anti-effect of ICI has not been established yet. In the methods paper published by Volk et al. used a model to mimic Epstein Baar virus (EBV)-associated post-transplant lymphoproliferative disorder (EBV-PTLD) by the infection of fully humanized NRG mice with EBV. They demonstrated that pembrolizumab induced expansion of EBV-positive tumor cells and infiltration of EBV-positive tumor cells into the central nervous system. This result is likely due to the emergence of dysfunctional T cells and may reflect clinical observations with spreading of EBV after ICI treatment.in vivo. In the mini-review paper by Sun et al., the authors summarize recent advances with HIS mice that carry autologous tumor cells. They specifically focus on BLT humanized mice engrafted with human fetal liver CD34+ cells transduced with a MLL-AF9-GFP retrovirus to assess the response to immunotherapies in human leukemia, in particular anti-CD19 CAR T-cells.The technical advances in the generation of humanized mice allow today to specifically investigate human immune cells Katano et al. report the generation of human IL-15 transgenic NOD/Shi-scid/IL-2Rγnull (NOG) mice, which lack the mouse Fcγ receptor and consequently do not produce antibody-dependent cellular cytotoxicity (ADCC). This allowed the authors to specifically examine the contribution of human NK cell-mediated ADCC to anti-tumor responses. Therefore, this new model may prove useful for the preclinical testing of antibody-based approaches known to induce ADCC.In their original article, Maser et al. studied the anti-tumor effect of plasmacytoid dendritic cells (pDCs) in NOG-hIL3/GM-CSFTg mice, which supported the development of human pDCs. In certain tumor models, pDCs were recruited to tumors growing in mice. These intratumoral pDCs were functional as they produced type I interferon (IFN) in response to Toll-like receptor stimulation, which may prove beneficial for promoting NK cell-mediated anti-tumor activity.Izumi et al. investigated the effect of an anti-CD64 dimeric antibody conjugated with pyrrolobenzodiazepine (dPBD) in vivo in NOG mice. They showed that the anti-CD64-dPBD antibody selectively depleted monocyte progenitors without affecting the mature compartment. This compound also prevented the development of chronic myelomonocytic leukemia in mice, a disease with high therapeutic need. Finally, they extended their work to solid tumors and showed that the anti-CD64-dPBD antibody led to tumor growth regression by eliminating the tumor-associated macrophage population.Based on the identification of human monocyte-restricted progenitors, Serr et al. summarizes the possible use of HIS mice to investigate and develop immunotherapies based on regulatory T-cells (Tregs). Tregs are known to facilitate tumor growth. The authors propose to use Treg inhibition to enhance anti-tumor responses in personalized humanized mouse models that are reconstituted with the patient’s immune system and tumor cells.The review by in vivo. In the review by Horowitz et al., the authors present humanized mouse models that can be used to study ILCs and potential therapeutic approaches that employ human ILCs against cancer. This review provides an excellent overview about the development of ILC-based cancer immunotherapy.Standard immunodeficient mice weakly support the development of human innate lymphoid cells (ILCs), in particular human NK-cells. Recent progress in the generation of humanized immunodeficient animals, in particular models that produce human IL-15, led to the development of human ILCs Flahou et al. review the use of HIS mice in the context of induced pluripotent stem cells (iPSC)-derived therapies for tissue regeneration. Here the authors suggest using humanized mouse models that develop human NK cells to evaluate the immunogenicity and rejection of transplanted iPSCs and their progeny. This could help to develop HLA-engineered iPSC-derived cells for universal clinical use.Gogoleva et al. review several humanized mouse models, which express human TNF in order to study human autoimmune diseases and the effect of TNF inhibitors on hematopoiesis in vivo.Tumor necrosis factor (TNF) is a pro-inflammatory cytokine that controls inflammation. TNF inhibitors have been widely used in the clinic to treat autoimmune or inflammatory diseases. Münz focuses on the evaluation of immune responses to control pathologies induced by human γ-herpesviruses in humanized mouse models and discusses strategies for developing vaccines.Human γ-herpesviruses such as Epstein Barr virus (EBV) and Kaposi sarcoma-associated herpesvirus (KSHV) can induce lymphoproliferative diseases in humans. The immune response against γ-herpesviruses has been largely recapitulated in humanized mouse models. The review article by Yong et al. tested two different immunotherapeutics, including human interleukin-2 (Proleukin) and the anti-CD3 monoclonal antibody OKT3, in a humanized NSG model. Administration of Proleukin to humanized mice resulted in an increase of several pro-inflammatory cytokines and chemokines. Likewise, the use of OKT3 treatment in their model led to T-cell depletion along with an increase in several pro-inflammatory cytokines. This indicates that the presented models can reproduce the effect of these immunotherapeutics observed in patients.Furthermore, This Research Topic shows the usefulness of HIS mouse models in translational research related to several important human diseases and we are looking forward to future developments in this exciting research area.All authors listed have made a substantial, direct, and intellectual contribution to the work, and approved it for publication. All authors contributed equally to the work.YS is supported by a Grant-in-Aid for Challenging Exploratory Research (20K21547) from the Ministry of Education, Culture, Sports, Science and Technology of Japan. TW is supported by a faculty-funded career position at Karolinska Institutet (2-1060/2018). APAT is supported by the Professor Dr. Max Cloëtta Foundation and the Swiss Cancer Research grant KFS-4875-08-2019.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Participants reported increases in knowledge level and confidence in offering STI testing and managing a greater variety of sexual health cases. This study demonstrates the feasibility of implementing a hub-and-spoke model to enable GPs to deliver sexual health care with support from a sexual health specialist service.Improving access to sexual health services is critical in light of rising sexually transmitted infections (STIs). We evaluated a hub-and-spoke model for improving access to sexual health services in three general practices in Victoria, Australia. The primary outcome was the impact on HIV and STI  testing. Segmented linear regression analysis was conducted to examine the trends in the total HIV/STI tests pre- (from January 2019 to June 2020) and post-implementation (from July 2020 to July 2021). We evaluated the feasibility and acceptability of integrating this model into the general practices using semi-structured individual interviews. There was a statistically significant rise in testing for HIV and STIs in all general practices: post-implementation, there was an increase of an average of 11.2 chlamydia tests per month ( Neisseria gonorrhoeae and Mycoplasma genitalium globally [The World Health Organization estimates more than a million new sexually transmitted infections (STIs) a day globally . In Austglobally .To control HIV/STI, healthcare systems must support earlier testing and management because a shorter duration of infectiousness has a powerful effect in reducing the incidence of HIV/STI . Most STGeneral practitioners play a critical role in improving HIV/STI testing access. In Australia, it is estimated that 83% of the general population would visit a general practitioner in a year . There hThe hub-and-spoke model is a network with a central facility that provides a full array of sexual health primary and specialist services (“the hub”) supporting geographically dispersed secondary services (“the spokes”), which provides primary but more limited specialist services . This arThis study aimed to evaluate a hub-and-spoke model for improving access and increasing HIV/STI testing in three general practices in Victoria, Australia. The primary outcome was to evaluate the impact on HIV and STI  testing. We assessed the feasibility and acceptability of integrating this model into the general practices using qualitative methods.In 2020, Melbourne Sexual Health Centre (MSHC), a part of Alfred Health, undertook a project funded by the Victorian Department of Health and Human Services in response to the Victorian Review of Sexual Health Services, which recommended the development of HIV/STI services provided in a decentralised hub-and-spoke model where Melbourne Sexual Health Centre served as the specialist hub for HIV/STI testing and treatment supporting GP spokes that provide primary care HIV/STI testing and treatment. The training phase of the project took place between May 2020 and July 2020. The support Melbourne Sexual Health Centre delivered to these three GP partner services is provided in the t-test to compare the average number of tests for each pathogen pre-implementation and implementation phase. Segmented linear regression analysis was conducted to examine the trends in the total HIV/STI tests in pre- (from January 2019 to June 2020) and post-implementation phases (from July 2020 to July 2021).We defined post-implementation in our study as after the month when clinics started seeing patients using the GP hub-and-spoke model. We extracted data from the pathology testing laboratory on the number of tests  and test positivity for each clinic from two time periods: (1) pre-implementation phase: January 2019 to June 2020; and (2) post-implementation phase: July 2020 to July 2021. Descriptive statistics were used to analyse the data. We used Student’s A total of 33 semi-structured individual interviews were conducted from May to June 2020, before the training: 5 from Clinic 1, 13 from Clinic 2, and 15 from Clinic 3. Interviews explored the staff’s current knowledge, attitude, and practice related to sexual health; current challenges in providing sexual health care in the general practice; and training needs. A second interview was conducted among staff who had experience providing the sexual health services in the respective clinics in August and September 2020. Interviews explored their experiences in delivering sexual health care as part of the hub-and-spoke model; feedback on the training received for the implementation of the sexual health service; changes in their knowledge, attitude, and practice as a result of participating in the training; and ongoing support needs. The interview guide is provided in the As this was part of a quality improvement and evaluation activity, we received a waiver from the Alfred Hospital Human Research Ethics Committee. A researcher (JO) approached each of the GP clinics’ practice managers to approach their clinic staff explaining the voluntary nature of the evaluation, and interviews were only conducted with those who agreed to participate. Consent was implied when people participated.p = 0.246, p = 0.026).p = 0.826, p = 0.001).Pre-implementation, there was no significant change in the mean monthly gonorrhoea tests .Pre-implementation, there was no significant change in the mean monthly syphilis tests .Pre-implementation, there was a significant decrease in HIV tests of 2.3 per month  is to increase STI testing coverage . Enablin18–2022 iThere are several advantages to the GP hub-and-spoke model. Strengthening general practices to provide sexual health care can help destigmatise sexual health services by providing them through GPs who are already accessible and trusted by the community. Patients who book in for a sexual health consultation with their GP may feel more comfortable discussing sexual health matters, knowing their GP is interested and skilled in providing this care . NormaliOur model of care uses the nurse to perform the initial assessment. This is beneficial for embedding STI testing in the practice, creating extra capacity for patient appointments, and workforce development for sexual health nurses . Our stuOur results must be read in light of some limitations. The evaluation was conducted during major disruptions to primary care services because of COVID-19-related social distancing restrictions. General practices during this period had to prioritise COVID-19-related activities (including delivering vaccines) and transition from in-person to telehealth services. Nevertheless, we still observed a significant rise in HIV/STI testing post-implementation. However, we did not observe any impact on test positivity, which could either reflect a lower incidence of STI because of declines in sexual activity  or an inThis study demonstrated that GPs are well placed to deliver sexual health care with specialist services providing support and training through a hub-and-spoke model. This ongoing and synergistic collaboration has led to a sustained increase in HIV and STI testing. Further implementation research is warranted to evaluate its scalability in other settings and for managing other conditions.


O-acyltransferase (GOAT), a member of the membrane-bound O-acyltransferase (MBOAT) enzyme family. The ghrelin/GOAT/GHS-R1a system is defined by multiple unique aspects within both protein biochemistry and endocrinology. Ghrelin serves as the only substrate for GOAT within the human proteome and, among the multiple hormones involved in energy homeostasis and metabolism such as insulin and leptin, acts as the only known hormone in circulation that directly stimulates appetite and hunger signalling. Advances in GOAT enzymology, structural modelling and inhibitor development have revolutionized our understanding of this enzyme and offered new tools for investigating ghrelin signalling at the molecular and organismal levels. In this review, we briefly summarize the current state of knowledge regarding ghrelin signalling and ghrelin/GOAT enzymology, discuss the GOAT structural model in the context of recently reported MBOAT enzyme superfamily member structures, and highlight the growing complement of GOAT inhibitors that offer options for both ghrelin signalling studies and therapeutic applications.The acylated peptide hormone ghrelin impacts a wide range of physiological processes but is most well known for controlling hunger and metabolic regulation. Ghrelin requires a unique posttranslational modification, serine octanoylation, to bind and activate signalling through its cognate GHS-R1a receptor. Ghrelin acylation is catalysed by ghrelin  Substitution at the acylation site serine of the truncated peptide with a diaminopropanoic acid (Dap) yields greater inhibition of GOAT at sub- to low-micromolar concentrations when using the hydrolytically stable octanoylated form, GSDap(C8:0)FL-[NH2]  . Permuta . PermutFL-[NH2] . These tin vitro inhibition of GOAT, there remained concerns whether potency can be maintained in a cellular or organismal context due to concerns with peptide cell permeability and biostability. A bisubstrate-mimetic analogue, GO-CoA-Tat, was developed in which the first 10-amino acids of ghrelin are linked with octanoyl-CoA through an amide bond and a cationic TAT sequence is appended to the end of ghrelin sequence to improve cell permeability [in vitro and cell assays without activating GHS-R1a receptor at sub- to low-micromolar concentrations [While early peptide-based ghrelin mimetics exhibited pronounced eability . GO-CoA-trations . Glucosetrations , and subtrations ,149. Whi50 values as low as 62 nM [In addition to the product and bisubstrate-mimetic inhibitors, we recently reported that ghrelin mimetic peptides incorporating an unmodified Dap residue in place of the acylation site serine act as potent GOAT inhibitors . These sas 62 nM . The pot. 7.2Many early GOAT inhibitors were peptide-based substrate or product mimetics that, except for GO-CoA-Tat, either lacked potency in cell-based assays or were not tested in these assays. Though GO-CoA-Tat exhibits promising results, as noted above biostability and potential for further inhibitor optimization remain a concern with peptidomimetics. Consequently, small-molecule GOAT inhibitors have been sought as more likely candidates for therapeutic development and deployment. Recent progress in this area brings hope that GOAT's value as a treatment avenue targeting ghrelin-dependent health conditions may be validated in the near future.The first reported small-molecule non-peptide-based GOAT inhibitors were synthesized and identified in 2011 from a library screen of compounds based on naphthalene analogues containing varying alkyl chains . These c50 = 8.4 µM. Structure-function analysis demonstrated the presence of an α-cyano-enone Michael acceptor is essential for inhibition and implicates the involvement of a nucleophilic residue, possibly cysteine, in the inhibition mechanism. Interestingly, inhibition was observed in the human isoform of GOAT but not in the highly similar mouse isoform . This contrast in inhibition profile suggests these inhibitors target a non-conserved residue and binding site in hGOAT, which would argue against direct engagement with the enzyme active site. Interestingly, one of the original CDDO derivative compounds found to inhibit GOAT, bardoxolone methyl, has been assessed as a therapeutic for inflammation and oxidative stress acting through the Nrf2-KEAP pathway. Clinical studies with bardoxolone methyl reported side-effects such as weight loss, reduction of diet-induced insulin resistance and improvements to glucose tolerance which are physiological effects that would be associated with alterations in the ghrelin-signalling pathway [Our research group screened a diverse small-molecule library and identified a set of synthetic triterpenoids that inhibit GOAT . These m pathway –156. Whiin vivo biostability and was determined to have an IC50 of 69 nM while exhibiting good PK/PD properties [et al. [in vivo studies were suggested. Wang reported another set of compounds as GOAT inhibitors based on a functionalized benzothiophene scaffold [et al. [Potent small-molecule GOAT inhibitors developed by pharmaceutical companies have recently begun to be reported in the scientific and patent literature. A set of compounds containing an aminopyrimidine scaffold which antagonize GOAT acylation have been reported to show inhibition activity in ELISA-based assays –161. Theoperties . This cooperties –165. Usi [et al.  identifiscaffold . Inhibitscaffold –170. God [et al. ,172 disc. 8As studies of ghrelin and its associated signalling pathway enter their third decade, the unique aspects of this peptide hormone and its impact across multiple physiological processes continue to motivate investigations. These studies span across the scientific spectrum from fundamental protein biochemistry to organismal metabolism and neuroendocrinology. At the molecular level, the most pressing challenges are the experimental determination of the structures of GOAT and the ghrelin•GHS-R1a signalling complex. Recent structures of the ghrelin receptor suggest the potential for rapid progress on this front, but the purification of enzymatically active GOAT remains a significant barrier to GOAT structural studies. The computational model of GOAT answered several questions regarding the enzyme architecture and revealed the transmembrane catalytic channel. Future studies by crystallography or cryo-EM will provide valuable insights into the substrate-binding sites and enzyme active site. We must also determine if GOAT itself is subject to regulation by posttranslational modifications through either direct impact on enzyme activity or effects on GOAT localization and trafficking across cellular membranes.Beyond expanding our understanding of the catalytic strategies employed by protein-modifying MBOAT family members, defining GOAT's structure will also inform inhibitor design and optimization. Potent and specific GOAT inhibitors are essential molecular tools for probing ghrelin's impact within the body as we move towards a complete model of ghrelin signalling at the organismal level. This model must address the biological role of des-acyl ghrelin, ghrelin reacylation at signalling target sites, the influence of ghrelin on behaviour and mental health, and all other aspects of ghrelin signalling within the body.Recognized at its discovery for its unique role in physiology and its potential as a therapeutic target, recent advances in the ghrelin/GOAT/GHS-R1a field have positioned us well to leverage opportunities to understand and harness ghrelin signalling. The creation of multiple potent GOAT inhibitors now enables the validation of ghrelin signalling as a treatment avenue for multiple metabolic disorders such as diabetes and obesity. At the molecular level, we can also now explore why ghrelin bears its unique octanoyl serine modification—perhaps as a mechanism for decoupling ghrelin signalling from the metabolic state of the ghrelin-producing cell by forcing the use of an exogenously derived acyl donor? The strong conservation of ghrelin and GOAT across the vertebrates also suggests an essential role for signalling, perhaps as a ‘survival hormone’ enhancing the ability of an organism to respond to multiple environmental stressors such as starvation over both acute and chronic time scales.Moving beyond metabolism and energy homeostasis, ghrelin can also provide a new avenue for understanding and treating addiction, stress and anxiety disorders, and other neurological conditions impacted by ghrelin signalling. The growing complement of molecular tools for modulating and blocking ghrelin signalling provides the ability to uncover and understand the involvement of this acylated hormone in a growing range of processes at the cellular and organismal level. Continuing efforts across the biochemical, biological and medical ghrelin research communities are essential for realizing these opportunities for defining and exploiting ghrelin signalling for therapeutic effect. The acceleration of progress across these fields in the last 5 years suggests we are entering an exciting new era in our understanding of ghrelin and its biological impact.


Long non-coding RNAs (lncRNAs) play an important role in genome regulation. Specifically, many lncRNAs interact with chromatin, recruit epigenetic complexes and in this way affect large-scale gene expression programs. However, the experimental data about lncRNA-chromatin interactions is still limited. The majority of experimental protocols do not provide any insight into the mechanics of lncRNA-based genome-wide epigenetic regulation. Here we present the HiMoRNA (Histone-Modifying RNA) database, a resource containing correlated lncRNA–epigenetic changes in specific genomic locations genome-wide. HiMoRNA integrates a large amount of multi-omics data to characterize the effects of lncRNA on epigenetic modifications and gene expression. The current release of HiMoRNA includes more than five million associations in humans for ten histone modifications in multiple genomic loci and 4145 lncRNAs. HiMoRNA provides a user-friendly interface to facilitate browsing, searching and retrieving of lncRNAs associated with epigenetic profiles of various chromatin loci. Analysis of the HiMoRNA data suggests that several lncRNA including JPX might be involved not only in regulation of XIST locus but also in direct establishment or maintenance of X-chromosome inactivation. We believe that HiMoRNA is a convenient and valuable resource that can provide valuable biological insights and greatly facilitate functional annotation of lncRNAs. Transcription in higher organisms is pervasive and produces a wide variety of noncoding RNAs , with loMany lncRNAs interact with chromatin and are essential for epigenetic control of specific genome loci as well as organization of the chromosomes ,11,12,13To address this problem computationally we developed an approach to search for lncRNAs whose presence in the cell is significantly correlated with the presence of epigenetic modification in a specific genome locus. Here we report the HiMoRNA (Histone-Modifying RNA) database, a resource containing coordinated lncRNA–epigenetic changes genome-wide. HiMoRNA integrates a large amount of multi-omics data to characterize the effects of lncRNA on epigenetic modifications and gene expression. The current release of HiMoRNA includes 5,640,250 associations in humans for the ten most studied histone modifications and 4145 lncRNA. HiMoRNA provides a user-friendly interface to facilitate browsing, searching and retrieving of lncRNAs associated with specific chromatin loci. Analysis of the HiMoRNA data suggests that several lncRNA, including JPX, might be involved not only in regulation of XIST locus but also in direct establishment or maintenance of X-chromosome inactivation. We believe that HiMoRNA is a convenient and valuable resource that can provide valuable biological insights and greatly facilitate functional annotation of lncRNAs.http://himorna.fbras.ru (accessed on 19 November 2021).Availability: https://www.ncbi.nlm.nih.gov/assembly/GCF_000001405.26/, accessed on 19 November 2021) and GENCODE gene annotation version 31 .All analyses were performed using human genome build hg38  for matching pairs of signal-control experiments for the same biosample. ChIP-seq peaks detected in all biosamples for one histone modification were merged using bedtools [We downloaded bam files  for ten histone modification marks  having the highest number of biosamples  from thebedtools  to standbedtools . We assobedtools  (SupplemWe downloaded RNA-seq data (gene counts) from the ENCODE portal for the same biosamples for which ChIP-seq data was used. Genes counts were normalized using DESeq2 separately for each group of RNA-seq samples matching ChIP-seq samples for every histone modification. As a list of lncRNAs we used gencode.v31.long_noncoding_RNAs and normalized the gene expression of each dataset with DESeq2. For each histone mark we further analyzed only a set of lncRNAs expressed in at least ten biosamples.To select lncRNAs potentially involved in epigenetics regulation of a specific locus across multiple cell types we calculated the Spearman correlation coefficient (SCC) between all lncRNAs expression vectors and all ChIP-Seq peaks over all samples available for every histone modification. Only significant correlations , target gene name, and SCC value. HiMoRNA also provides a download button allowing users to retrieve all of the interaction data.To further explore the interaction of interest, users can click on a histone modification, an lncRNA, a gene or a correlation in the table to launch a new page that contains the corresponding metadata and visualizations. LncRNA page provides information about its expression in all cell types used in HiMoRNA, per chromosome distribution of correlated peaks for each histone modification separately and a summary table of all peaks correlated with this lncRNA. The histone modification page provides information about chromosomal distribution of ChIP-seq peaks and a table of all lncRNA correlated with at least one peak of this modification. The gene page provides information about its expression in all cell types used in HiMoRNA, a table of all histone modification peaks associated with the gene and a summary table of all peaks associated with this gene and at the same time correlated with any lncRNA. The correlation page provides detailed information about how this correlation has been obtained, including lncRNA expression and ChIP-seq peak signal values in all the cell types used to calculate SCC.LncRNA-locus correlations and the results of multi-omics data set analysis were stored and queried by neo4j. The web interface was implemented in JavaScript and Vue and has been tested on Firefox, Chrome and Safari.https://github.com/lab-medvedeva/himorna-frontend (accessed on 19 November 2021) (platform frontend) and https://github.com/lab-medvedeva/himorna-backend (accessed on 19 November 2021) (platform backend).Source code and database dumps in different formats are available at HiMoRNA provides a comprehensive collection of lncRNA–genomic loci for which lncRNA expression is significantly correlated with the histone modification signal across multiple cell and tissue types. HiMoRNA currently contains over five million correlations for ten histone modifications and 4145 lncRNA in humans. For each lncRNA, histone modification and gene we summarized the metadata to facilitate the study of lncRNAs function in epigenetic regulation. HiMoRNA also provides a user-friendly web interface for customized searching, browsing and accessing the interaction data.Deeper investigation of per chromosome distribution of ChIP-seq peaks associated with specific lncRNAs demonstrated significant nonuniformity. For seven out of ten studied histone modifications, at least one lncRNA demonstrated an enrichment of correlated ChIP-seq peaks at one of the chromosomes. The most striking cases include lncRNA JPX and TSIX for which the X-chromosome contains the majority of the correlated peaks for H3K9me3 and H3K27me3, respectively , while tX-chromosome inactivation is an essential epigenetic process which is regulated by several lncRNAs, including XIST, the master regulator of X-inactivation initiation. Other lncRNAs have also been implicated in regulating XIST, including TSIX, JPX, and FTX. Tian et al.  proposedIt has been found recently that Jpx acts as a CTCF release factor and in this way affects a 3D structure of the genome by regulating anchor site usage . CTCF isFinally, we observed several other lncRNAs significantly correlated with the ChIP-seq peaks of a particular histone modification on the X-chromosome (ST3) for which experimental evidence of participation in X-chromosome inactivation is still required.All the above suggest a possible mechanistic explanation of the role of several lncRNAs in establishing and maintaining the epigenetic profile of inactive X-chromosome. We realize that correlation of the ChIP-seq peaks and lncRNA expression does not guarantee causal relationship between these two events. Still, we hypothesize that at least some of these lncRNA could participate in epigenetic complex formation or attract them to specific X-chromosome loci. This example shows nicely how HiMoRNA enables researchers to translate associations between lncRNA and histone modifications into novel biologic insights.The area of functional lncRNA annotation is moving fast. We are fully committed to the maintenance and update of HiMoRNA, and we will monitor new data on the role of lncRNA in epigenetic regulation and expand the database as data becomes available. Specifically, we will add experimental data on lncRNA-chromatin interactions available databases to support our predicted lncRNA-genomic loci interactions. We also will incorporate data on lncRNA knockdown effects of gene expression from FANTOM6 . As more


Segmental colitis associated with diverticulosis (SCAD) is an inflammatory disease affecting segments of the large bowel with diverticular disease. SCAD presents several challenges in diagnoses and treatment because it often mimics a range of disorders including inflammatory bowel disease and malignancy. Here, we present the case of a 72-year-old man with lower abdominal pain and bloody stools whose initial abdominal workup showed nonspecific large bowel thickening and concerns for malignancy. Ultimately, the patient was diagnosed with mild SCAD and treated conservatively with a resolution of symptoms. He had no symptoms at the three-month and 1-year follow-ups. This case highlights the importance of including SCAD in the initial differential diagnosis to allow accurate identification and treatment. Segmental colitis associated with diverticulosis (SCAD) is an inflammatory disease affecting segments of the large bowel with diverticular disease -3. SCAD A 72-year-old man with a medical history of glaucoma, hyperlipidemia, and diabetes mellitus type 2 was evaluated for a one-day history of left lower abdominal pain and intermittent bloody stools. His complaints were associated with progressive abdominal pressure and discomfort for the same duration. The patient’s vitals were remarkable for a blood pressure of 150/84 mmHg, normal heart rate, and temperature. Laboratory testing including complete blood count and biochemistry were unremarkable. Computed tomography (CT) of the abdomen and pelvis revealed an abnormal thickening of the sigmoid colon concerning for acute sigmoid diverticulitis or malignancy Figure . Follow-SCAD is an inflammatory disease affecting segments of the large bowel with a diverticular disease that typically involves the sigmoid colon and can mimic a malignancy or IBD. It can present with a wide range of symptoms such as rectal bleeding, diarrhea, and abdominal pain. Segmental colitis (or diverticular colitis) is commonly associated with varying complications such as bleeding, perforation, and formation of abscesses . The terVisualization of the involved mucosa further allows for the subclassification of SCAD into one of four types (type A to D) by histologic appearance. Histology ranges from crescentic fold pattern (type A) to severe ulcerative colitis-like pattern (type D) -9. HowevThis case highlights the need to include SCAD as part of the differential diagnosis. Treatment of SCAD is not clearly defined. Therapy is geared toward reducing mucosal inflammation in moderate-to-severe SCAD with mesalamine, corticosteroids, and antibiotics . PhysiciSCAD can present with a broad spectrum of symptoms. Although visualization of the involved mucosa is the best means for diagnosing the disease, care must be taken to avoid misclassification based on endoscopic or radiographic findings including colonic thickening. Therefore, physicians must maintain a high suspicion for SCAD as a possible explanation for abnormal colonic thickening.


Scientific Reports 10.1038/s41598-020-67411-w, published online 03 July 2020Correction to: The original version of this Article contained an error in the Abstract.“Preclinical models that reliably recapitulate the immunosuppressive properties of human gliomas are essential to assess immune-based therapies. GL261 murine glioma cells are widely used as a syngeneic animal model of glioma, however, it has become common practice to transfect these cells with luciferase for fluorescent tumor tracking. The aim of this study was to compare the survival of mice injected with fluorescent or non-fluorescent GL261 cells and characterize the differences in their tumor microenvironment.”now reads:“Preclinical models that reliably recapitulate the immunosuppressive properties of human gliomas are essential to assess immune-based therapies. GL261 murine glioma cells are widely used as a syngeneic animal model of glioma, however, it has become common practice to transfect these cells with luciferase for bioluminescent tumor tracking. The aim of this study was to compare the survival of mice injected with bioluminescent or non-bioluminescent GL261 cells and characterize the differences in their tumor microenvironment.”The original Article has been corrected.


Several studies began also to illustrate that trans-acting RNA modulators can adjust the timing and efficiency of frameshifting illuminating that frameshifting can be a dynamically regulated process in cells. Here, we intend to summarize these new findings and emphasize how it fits in our current understanding of PRF mechanisms as previously described.Translation facilitates the transfer of the genetic information stored in the genome via messenger RNAs to a functional protein and is therefore one of the most fundamental cellular processes. Programmed ribosomal frameshifting is a ubiquitous alternative translation event that is extensively used by viruses to regulate gene expression from overlapping open reading frames in a controlled manner. Recent technical advances in the translation field enabled the identification of precise mechanisms as to how and when ribosomes change the reading frame on mRNAs containing  The incorporation of the correct amino acid is facilitated by the match between the mRNA codon and its cognate anticodon of the tRNA that delivers the appropriate amino acid. The genetic code is universal and read in triplets directed from the mRNA 5′ to 3′ end. The movement of tRNAs and the mRNA through the ribosome is maintained by coordinated, inter- and intra-subunit conformational changes and rotations of the ribosome . During nslation . An intenslation  some of nslation . Translanslation . While tnslation , in thisnslation .Escherichia coli dnaX gene   . Movemen(PEG-10) . –1PRF, (PEG-10)  and –2PR(PEG-10)  have bee–1PRF is extensively studied in RNA viruses, including the coronaviruses   and retrcis-acting elements in the mRNA that are separated by a spacer sequence: a heptanucleotide slippery sequence on which the ribosome can slip into the alternative frame   , where tProtein-mediated –1 frameshifting has been more recently discovered on the cellular poly-(C) binding proteins (PCBP) that promote –1 as well as –2 frameshifting in arteriviruses such as the PRRSV by directly interacting with the viral nonstructural protein nsp1β . In additrans-factor-mediated translational regulation has been discovered in the cardioviruses Encephalomyocarditis Virus (EMCV) and Theiler’s Murine Encephalomyelitis Virus (TMEV) (Another elegant example of a pathogen-encoded s (TMEV) . The cods (TMEV) . Howevers (TMEV) , the dows (TMEV) . In addis (TMEV) . Furthers (TMEV) . The incRecent work combining structural, biochemical and single-molecule analysis illuminated how the unique RNA binding fold found in the 2A structure allows it to interact with translating ribosomes and the downstream RNA element  Figure . The 2A Such viral-encoded factors seem to selectively recognize particular RNA motifs and stabilize the RNA to impede translational elongation. Pausing at these sites likely opens a favorable time-window for codon-anticodon interactions to be re-established in the alternative reading frame, however, exactly which step of translation elongation is affected is still an open question.in vitro  protein is associated with the IBV frameshift RNA element in vitro . InteresPEG-10 mRNA   and the -10 mRNA . In summtrans-acting factor directly influencing –1PRF during SARS-CoV-2 infections , micro RNAs (miRNAs)] have already been successfully used for in vitro studies on frameshifting and consequently be suggested to act as potential effectors of the frameshifting process  ribosome undergoes large conformational changes facilitating the translocation of the tRNAs from the P- and A-sites into the E- and P-sites, along the mRNA in one codon steps  Figure . Since rng frame . The codng frame   also suggested that timing of +1 frameshifting could be similar to canonical –1 frameshifting occurring during late steps of translocation (Recent studies employing a +1 frameshift-prone mRNA and native location . Structulocation . In thislocation . This polocation . Nonethelocation . Whethercis-acting canonical frameshift elements. Also, discovery of novel cellular and viral trans-acting factors, nascent polypeptide chain interactions and modifications on the RNAs affecting frameshifting, continuously reveal new levels of complexity. In the long run, the accumulated knowledge about this recoding mechanism and its regulation in cells will help to pave the way for therapeutic studies inhibiting frameshifting on pathogen mRNAs. Potential antiviral therapeutics could be designed to bind the frameshift RNA in a way that the stimulatory secondary structure either cannot be formed or cannot be unwound, hence prohibiting viral protein synthesis. In this respect, small synthetic, complementary oligonucleotides have already been suggested as possible therapeutic agents (Despite fundamental knowledge gained about frameshifting events over the past years, there are still many outstanding questions concerning the detailed molecular mechanisms and occurrences of PRF. For instance, both upstream and downstream regions of frameshift RNAs seem to influence folding and the function of c agents . Artificc agents . Additioc agents , doxorubc agents , naphthyc agents  or merafc agents  are desi


Gene mapping indicated that the mutant was controlled by a gene located on chromosome nine. Phenotypic and cytological observations revealed that dnl2 showed inhibited cell growth, altered vascular bundle patterning, and disrupted secondary cell wall structure when compared with the wild-type, which could be the direct cause of the dwarf and narrow-leaf phenotype. The phytohormone levels, especially auxin and gibberellin, were significantly decreased in dnl2 compared to the wild-type plants. Transcriptome profiling of the internodes of the dnl2 mutant and wild-type revealed a large number of differentially expressed genes enriched in the cell wall biosynthesis, remodeling, and hormone biosynthesis and signaling pathways. Therefore, we suggest that crosstalk between hormones  may contribute to the dwarf and narrow-leaf phenotype by influencing cell growth. These results provide a foundation for DNL2 gene cloning and further elucidation of the molecular mechanism of the regulation of plant height and leaf shape in maize.Lodging is the primary factor limiting high yield under a high plant density. However, an optimal plant height and leaf shape can effectively decrease the lodging risk. Here we studied an ethyl methanesulfonate (EMS)-induced dwarf and a narrow-leaf mutant,  Zea mays L.) is one of the most important cereal crops in the world. Studies have demonstrated that increasing the planting density is an essential approach in order to increase per-hectare yield potential in maize  directly. The read count numbers of fragments per kilobases per million reads (FPKM) were converted using Stringtie v2.1.0 software. The differential expression analysis between the wild-type and the dnl2 mutant was performed was performed using DESeq2. The resulting p-values were adjusted using the Benjamini and Hochberg’s approach for controlling the false discovery rate (FDR). Genes with Log2 fold-change (Log2FC) ≥ 1 (up-regulated) or Log2FC ≤ −1 (down-regulated) and FDR < 0.01 were considered as differentially expressed genes (DEGs).After removing reads containing adapters or poly-N and low-quality reads  pathways.GO enrichment analysis of the DEGs was implemented using the GOseq R package and GO terms with corrected tubulin was used as an internal control in the qRT-PCR. The reaction was performed in a 96-well plate on a CFX96 real-time PCR detection system  using TB Green II Premix Ex Taq  using the thermal cycling parameters . The relative expression level of the selected DEGs was calculated with the 2−ΔΔCT method [The expression levels of some DEGs were evaluated by qRT-PCR to validate the RNA-seq data. The specific primers for qRT-PCR are provided in T method . The readnl2, which exhibited short internodes, narrow leaves, and various developmental defects. Genetic analysis suggested the DNL2 was located near the centromere of chromosome nine. The phenotypic, cytological, and biochemical comparison between the dnl2 and the wild-type mutants revealed that the cell growth, vascular bundle patterning, cell wall structure, and phytohormone contents were altered in the internodes and leaves of dnl2, which were the main causes of the defective phenotype. The transcriptome analysis further proved that the crucial genes involved in cell wall development, phytohormone synthesis, and signaling were differentially expressed between the dnl2 and wild-type plants. Our study provides important clues for the further elucidation of the molecular mechanism of the regulation of plant height and leaf shape in maize.In this study, we characterized a recessive maize mutant,


The redox status of a host cell is an important factor in viral entry due to the unique conditions required for the conformational changes that ensure the binding and entry of a virus into the host cell. Upon entry into the airways, viral replication occurs and the innate immune system responds by activating macrophage and dendritic cells which contribute to inflammation. This review examines available literature and proposes mechanisms by which oxidative stress and inflammation could contribute to COVID-19 pathogenesis. Further, certain antioxidants currently undergoing some form of trial in COVID-19 patients and the corresponding required research gaps are highlighted to show how targeting oxidative stress and inflammation could ameliorate COVID-19 severity.COVID-19 is a zoonotic disease with devastating economic and public health impacts globally. Being a novel disease, current research is focused on a clearer understanding of the mechanisms involved in its pathogenesis and viable therapeutic strategies. Oxidative stress and inflammation are intertwined processes that play roles in disease progression and response to therapy  The coronavirus disease 2019 (COVID-19) is a multi-organ disease caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). In March 2020, the World Health Organization (WHO) declared the coronavirus disease (COVID-19) a pandemic with about 571,678 confirmed cases and 26,494 deaths declared globally at that time . Many coSARS-CoV-2 is enveloped virus with a single positive-stranded RNA genome of approximately 26-32 kb . The corA combination of factors comes into play to make viral entry into a host cell and subsequent replication in the host possible . A numbe2+ can result in the production of other radicals like hydrogen peroxide (H2O2), hydroxyl radical (OH•), and organic peroxides. The mitochondrial respiratory chain also produces nitric oxide (NO), which can generate other reactive nitrogen species  overwhelms the cellular antioxidant buffering capacity, eventually resulting in damage to cellular macromolecules . The pro species .Reactive oxygen species play a number of important roles in normal cell physiology. In endothelial cells, angiotensin II stimulates  oxidase . Downstrferation . In the pressure . The rolpressure . Macrophpressure . The cytpressure . These eOn the other hand, the deleterious effects of reactive species can be seen in the oxidation of cellular macromolecules like lipids, proteins and carbohydrates resulting in alterations in their functions. The hydroxyl radical, which is highly reactive with a short half-life can react with DNA bases to form adducts that can alter transcription resulting in altered protein function . Peroxylc and eventually death of the cell  and 4-hydroxynonenal that affect the compactness and integrity of cell membranes . Oxidatithe cell . Lipids the cell . This althe cell . A crossthe cell .Oxidation of proteins, especially those rich in cysteine-residues results in modifications that cause protein aggregation and altered function due to negative metabolic impacts on protein’s three dimensional structure . The hydvia mechanisms independent of IκB phosphorylation  which is vital in host entry, amplification and subsequent pathogenesis of SARS-CoV-2 can also contribute to oxidative stress in the pathogenesis of the disease . ACE-2 iTo protect the body against pathogens, cells have the unique ability to identify and restrict the replication of pathogens. The immune response can be broadly divided into the innate immune system and adaptive immune system  with innin silico molecular docking study which showed significant binding between the spike protein of SARS-CoV-2 and toll-like receptors especially TLR4 (Toll-like receptors (TLRs) are a prominent class of PRRs, existing as integral glycoproteins with an extracellular luminal ligand binding domain and a cytoplasmic signaling receptor homology domain . Once a lly TLR4 .A limitation of the recognition mechanism of immune response is that pathogens can sometimes escape recognition once they alter an identifying molecular feature like the order of monomers in its polymers . AnotherThe immune response is also characterized by assembly and activation of inflammasomes induced by ROS production . The actThe cross talk between oxidative stress and activation of pro-inflammatory cytokines contributes to damage of cellular macromolecules and tissues observed in COVID-19 patients Figure 12+ which reacts with molecular oxygen and hydrogen peroxide to generate superoxide anion and hydroxyl radical respectively  for instance, the possible therapeutic effects of vitamin C in COVID-19 therapy has received attention . StudiesVitamin D comprises a number of fat-soluble steroids with anti-inflammatory and immune modulatory properties, for which 1,25-dihydroxyvitamin D is the active form . The infvia interactions with a number of cellular proteins, signaling molecules and enzymes  is an immediate precursor of glutathione . GlutathCurcuma longa also known as turmeric (In silico docking results have shown that the viral spike protein and ACE-2 possess binding affinity to curcumin via covalent and non-covalent interactions, suggesting that the compound can interfere with SARS-CoV-2 entry into host cells (Curcumin, a hydrophobic polyphenol, is the active constituent of turmeric . Curcumiturmeric . In silist cells . Considest cells . An ongost cells in vitro and in vivo models of diseases for which oxidative stress and inflammation are implicated (Quercetin is a plant flavonoid with reported antioxidant, anti-inflammatory and antiviral effects . Althougplicated . HoweverOxidative stress and inflammation are interconnected processes that contribute to COVID-19 progression and response to therapy. Natural antioxidants can counteract altered signaling pathways activated during COVID-19 pathogenesis. The highlighted research gaps show that further investigations are needed along this line to provide efficient counteracting strategies to ameliorate severity of the disease and improve treatment outcomes especially in patients with underlying complications.GF - conception and drafting of manuscript. AM - research and drafting of manuscript. OO - research and drafting of manuscript. SM - research and drafting of manuscript. CM - conception and drafting of manuscript. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.