Objective: This study aims to identify an immune-related signature to predict clinical outcomes of oral squamous cell carcinoma (OSCC) patients. Methods: Gene transcriptome data of both tumor and normal tissues from OSCC and the corresponding clinical information were downloaded from The Cancer Genome Atlas (TCGA). Tumor Immune Estimation Resource algorithm (ESTIMATE) was used to calculate the immune/stromal-related scores. The immune/stromal scores and associated clinical characteristics of OSCC patients were evaluated. Univariate Cox proportional hazards regression analyses, least absolute shrinkage, and selection operator (LASSO) and receiver operating characteristic (ROC) curve analyses were performed to assess the prognostic prediction capacity. Gene Set Enrichment Analysis (GSEA) and Gene Ontology (GO) function annotation were used to analysis the functions of TME-related genes. Results: Eleven predictor genes were identified in the immune-related signature and overall survival (OS) in the high-risk group was significantly shorter than in the low-risk group. An ROC analysis showed the TME-related signature could predict the total OS of OSCC patients. Moreover, GSEA and GO function annotation proved that immunity and immune-related pathways were mainly enriched in the high-risk group. Conclusions: We identified an immune-related signature that was closely correlated with the prognosis and immune response of OSCC patients. This signature may have important implications for improving the clinical survival rate of OSCC patients and provide a potential strategy for cancer immunotherapy. Oral squamous cell carcinoma (OSCC) is one of the most common malignancies arising in the oral cavity worldwide, and the incidence of OSCC has been increasing by at least 1% annually, especially in China . Common Immunotherapy is one of the recent breakthroughs in cancer therapy and becoming a new promising approach in cancer treatment, including OSCC ,8. HowevImmune and stromal cells are two most abundant cell types and their degree of infiltration in the TME has a clinical prognostic value in many cancer types ,16,17. TAC103563.1, CCL22 (C-C motif chemokine ligand 22), FLT3 (FMS-like tyrosine 3), GALR2 , IGKV1D.8, IGLV1.36, IGLV4.60, IL10 (interleukin 10), LINC00861, LINC01508, and MS4A2  was constructed and confirmed to be significantly associated with the overall survival (OS) of OSCC patients. In conclusion, we screened the immune-related signature and explored the relationship between the screened immune-related signature and the prognosis of OSCC patients, and we aimed to use this signature as a potential prognostic biomarker and immune therapeutic target for OSCC patients.In this study, the stromal and immune scores of 319 patients from the TCGA OSCC database was determined using the ESTIMATE algorithm. A total of 562 upregulated and 31 downregulated immune-related genes were identified. Moreover, a functional enrichment analysis showed that the immune-related genes mainly played a critical role in immune response, activation/proliferation of immune-related cells, and chemokine activity. Finally, a prognostic 11 genes model, i.e., n = 319) was obtained from the National Cancer Institute GDC Data Portal  (accessed on 3 December 2020). Patients with complete follow-up data and survival status information were included in this analysis. Immune and stromal scores were calculated using the ESTIMATE algorithm [The RNA-sequence dataset  was used to screen for differentially expressed genes (DEGs) between OSCC tissue and normal tissues. FDR < 0.05 and |log2FoldChange| > 1 was considered as threshold values for the identification of DEGs. A heatmap was used to visualize the expression profiles of the DEGs. Venn diagrams were used to show the overlaps among the DEGs .https://string-db.org) (accessed on 3 December 2020) was used to predict the functional interaction of proteins. Cytoscape software was used to analyze the interaction scores of the PPI network nodes, and scores > 0.95 were considered the key PPI nodes [Gene Ontology (GO), including biological processes (BPs), molecular functions (MFs), and cellular components (CCs) were determined using the R package “ggplot2”, “clusterProfiler” and “enrichplot”, respectively. The Kyoto Encyclopedia of Genes and Genomes (KEGG) was used for the enrichment analysis of pathways . The SeaPI nodes .n = 319) were divided into high-risk and low-risk groups based on immune and stromal scores estimated by ESTIMATE. The Cox proportional hazards regression model was used to predict the OS of OSCC patients. p < 0.05 was statistically significant. Univariate Cox proportional hazards regression analyses were used to analyze the survival-related DEGs. The survival-related DEGs were put into the Cox proportional hazards model survival analysis with a least absolute shrinkage and a selection operator (LASSO) penalty. Finally, the TME-related prognostic signature was constructed by weighting the Cox regression coefficients to calculate a risk score for each patient. Using the cut-off value with the median value of the risk score, patients were classified as low-risk and high-risk.OSCC patients  was used to perform the statistical analysis. The R code used in this study was showed in Text S1.Univariate and multivariate Cox proportional hazards regression analyses were performed using the R package “survival”. A LASSO Cox survival analysis was performed using the R package “glmnet”, and a 1000-fold cross-validation was used. The R package “time-ROC” and “time AUC” were used to determine the ROC curves and area under the curve (AUC) for the predictive ability of the TME-related signatures. A heatmap was used to visualize the expression profiles of the DEGs. The alluvial diagrams were analyzed using “ggalluvial” in R. The clinical data of 319 patients with OSCC downloaded from the OSCC-TCGA RNA-seq database were analyzed in the present study. To explore the relationship between stromal and immune scores and the clinical characteristics of OSCC patients, a total of 19,467 genes were extracted from the OSCC-TCGA RNA-seq database. It is well-known that stromal and immune cells are two main types of nontumor components in the TME and have been identified to be of value for the diagnosis and prognostic evaluation of tumor. Therefore, the stromal and immune scores of 319 OSCC patients were determined using the ESTIMATE algorithm, and higher immune and stromal scores in the TME resulted in more stromal and immune components in the TME. Furthermore, the ESTIMATE score is the sum of the immune score and the stromal score and represents the combined ratio of the two components in the TME. Thus, these three scores represent the proportion of stromal cells, the proportion of immune cells, and the purity of tumor in the immune microenvironment, respectively ,27. In tIn order to determine the exact changes in gene expression profiles related to immune and stromal components in TME, we investigated the TME-related differentially expressed genes (DEGs) with high and low immune and stromal scores in OSCC patients with the R package “limma”. A total of 1625 DEGs were obtained from the immune score (high and low score samples), with 1227 genes upregulated and 398 genes downregulated a,c,d. SiTo explore the biological functions of the above TME-related DEGs, GO and KEGG enrichment analyses were performed using the R package “cluster Profiler”. As shown in ATP8B4, CD53, LILRB2, ITGAX, CD3G, IGHV3-11, SUCNR1, P2RY13, GPR183, GALR2, FPR3, CCR8, CCL13, C5AR1, ADORA3, P2RY12, CXCR3, CX3CR1, CCR4, ADRA2A, CCR1, FPR1, CCR2, CCR5, ITGB2, FCER1G, C3AR1, C3, and FPR2  database to construct a PPI network. In our study, CYTOSCAPE software was used to construct a PPI network based on the STRING database, and the interaction between 593 DEGs in . The topand FPR2 .AC093278.2, AC103563.1, AL365361.1, BTLA, CCL22, CCR4, CD5, CELF2, F5, FCRL3, FLT3, GALR2, IGKV1D-8, IGLV1-36, IGLV4-60, IL10, LINC00861, LINC01508, MS4A2, P2RY14, P2RY8, PPP1R16B, and RUBCNL (AC103563.1) − 0.0289 × (expression of CCL22) − 0.2829 × (expression of FLT3) + 0.03681 × (expression of GALR2) + 0.0048 × (expression of IGKV1D.8) + 0.0067 × (expression of IGLV1.36) − 0.0139 × (expression of IGLV4.60) − 0.0456 × (expression of IL10) − 0.2040 × (expression of LINC00861) + 0.1421 × (expression of LINC01508) − 0.0245 × (expression of MS4A2). The median value was taken as the cutoff, the OSCC patients with higher risk scores than the median value were classified in the high-risk group, and the OSCC patients with lower risk scores than the median value were classified in the low-risk group.To analyze the survival-related immune-DGEs, univariate Cox proportional hazards regression model survival analyses were performed, and a total of 23 prognostic genes were identified as survival-related DEGs: d RUBCNL . The lead RUBCNL a,b. FinaTo test the effect of the above model on the evaluation of OSCC patients in the future, we analyzed the outcome survival (OS) of the high-risk group and the low-risk group of OSCC patients. The risk distribution, survival status, and gene expression pattern are shown in To verify the clinical value of the 11-gene TME-related signature for predicting OS in OSCC patients, we used an external GEO-OSCC cohort (GSE65858) to validate our study. The 11-gene TME-related signature was constructed in GSE65858, and the risk scores were analyzed by the same methods. High- and low-risk OSCC groups were classified according to the median risk score. A Kaplan-Meier curve analysis showed that the low-risk score was closely associated with a longer overall survival time d, which In the present study, differentially expressed immune-related genes in oral squamous carcinoma cancer (OSCC) samples were identified and were used to construct a signature with 11 immune-related genes, which performed well in predicting the outcomes of OSCC patients, respectively.OSCC ranks as the eighth most common form of oral malignancy, with an estimated 500,000 new cases being reported annually, and OSCC is associated with a high grade, rapid progression, poor treatment effects, and bad outcomes . The cliThe TME is the cellular environment in which tumor cells live, which consists of an extracellular matrix, soluble molecules, and tumor stromal cells . The TMEIn the present study, we conducted a comprehensive analysis between the immune cells and immune-related genes and the clinical indexes and outcomes of OSCC patients. We found the following findings: (1) The clinical indexes  of OSCC patients were associated with the stromal, immune, and ESTIMATE scores, but the correlation was not significant . HoweverThe clinical implications of these 11 key genes have been reported in different types of cancers. For instance, CCL22 (C-C motif chemokine 22) controls T cell immunity by recruiting regulatory T cells to the tumor tissue and promoting regulatory T cell communication with dendritic cells in TME . Higher Similar studies have also been shown in OSCC. For instance, Li et al. selected eleven immunosuppression genes (ISGs), including INHBB, BGLAP, CTLA4, CALCA, CXCL8, IL22, FGFR3, HPRT1, ORMDL3, SPHK1, and TLR3, which showed a prognostic potential for OSCC . In thisHowever, our study still has limitations. For example, the OSCC TME-related signature was not verified by biological experiments both in vivo and in vitro, but in our ongoing projects, these immune-related genes and their potential mechanisms in OSCC are being verified. Moreover, due to the limited samples in the OSCC subgroups, the prognostic value of the immune-related signature in some OSCC subgroups did not show any statistical significance. Therefore, a large cohort of OSCC patient samples is needed for future validations. In summary, we proposed a signature of 11 immune-related genes based on the TME in OSCC, which could be used as an independent prognostic biomarker for OSCC patients. The signature of 11 immune-related genes had a predominant performance in obtaining the prognosis of OSCC patients, and it could achieve a more personalized and precise immunotherapy effect in OSCC.


Therefore, this review provides compelling evidence that SRPX2 might be a therapeutic target for inflammation and cancer-related inflammation for future cancer therapeutics.Accumulating evidence has showed that sushi-repeat-containing protein X-linked 2 (SRPX2) is an abnormal expression in a variety of cancers and involved in cancer carcinogenesis, chemosensitivity, and prognosis, which mainly promote cancer cell metastasis, invasion, and migration by regulating the uPAR/integrins/FAK signaling pathway, epithelial-mesenchymal transition (EMT), angiogenesis, and glycosylation. Inflammation has been regarded as a key role in regulating cancer initiation, progression, EMT, and therapeutics. Furthermore, SRPX2 exhibited excellent antifibrosis effect  E2A-HLF fusion gene in t -positive leukemia cells. It is also called as sushi-repeat protein upregulated in leukemia (SRPUL) [SRPX2, SRPX , SVEP1 (selectin-like protein), SELE (selectin E precursor), and SELP (selectin P precursor) [β1, NFATC3, MAN1 (LEM) domain containing 1 (LEMD1), miR-192, miR-215, and miR-149 [Sushi-repeat-containing protein X-linked 2 (SRPX2) is first found in 1999 by Kurosawa et al. that it is a downstream molecule of the ecursor) . As a crecursor) , 5. SRPXecursor) , 7. Rollecursor) –10. In a miR-149 –14.via TGFβR1/SMAD3/SRPX2/AP1/SMAD7-positive feedback loop, indicating that SRPX2 might be a therapeutic target for inflammation and cancer-related inflammation for future cancer therapeutics [SRPX2 is an abnormal expression in a variety of cancers, such as pancreatic cancer, colorectal cancer, and gastric cancer –17. Its apeutics . TherefoSRPX2 gene encodes a secreted extracellular matrix protein SRPX2 (GenBank NP_055282), which was identified as a chondroitin sulfate proteoglycan later [On chromosome Xq22.1, the an later . SRPX2 pan later . A sushian later , 25. Stuan later . Moreovean later .SRPX2 mRNA in the placenta, lung, trachea, uterus, and adrenal gland were high expression, whereas the levels were relatively low in the peripheral blood, brain, and bone marrow by detecting 24 normal human tissues with real-time RT-PCR [SRPX2 gene and its functions in cancer, the mRNA expression from the TCGA and GTEx database (including 29 types of cancer) was analyzed, and SRPX2 was found to abnormally express in multiple tumors  is a severe lung disease. Its progression is closely related with oxidative stress, mitochondrial dysfunction, and endoplasmic reticulum stress . Fibrobluced FMT . These fThis process of cancer spread is known as metastasis. A major process of metastasis is the proteolytic degradation of the extracellular matrix (ECM) to promote tumor cell migration and invasion . Currentβ-catenin pathway [β-catenin pathway-mediated MMP2/9 expression [FAK is a nonreceptor tyrosine kinase and aggregates to the cytoplasmic tails of integrins to regulate cellular adhesion, migration, invasion, and metastasis by activating a series of cellular signaling pathways such as the matrix metalloproteinases (MMPs), PI3K/Akt, and Wnt/ pathway –33. Rese pathway , 34. In  pathway . Moreovepression . SRPX2 hpression . Therefoin vivo. Further study showed SRPX2 knockdown increased the phosphorylation of Yes-associated protein (YAP), thus decreasing nuclear translocation to activate the Hippo signaling pathway, suggesting that SRPX2 might activate the Hippo signaling pathway to increase cell invasion in osteosarcoma [via the EMT process by mediating the PI3K/Akt/mTOR signaling pathway [β-catenin signaling pathway [β-catenin signaling pathway.EMT is a highly dynamic process characterized by transformation of polarized epithelial cells into highly invasive mesenchymal cells, resulting in the increased capacity of migration and invasion . The eveosarcoma . It was  pathway . In glio pathway . In addi pathway . TherefoAngiogenesis is the process through which novel blood vessels are formed from the existing blood vessels, and provides sufficient supply of nutrients and oxygen to cells. Tumor angiogenesis is closely related with tumor growth, progression, and metastasis . MiljkovO-glycans in healthy tissues, cancer-associated O-glycans are often truncated [COSMC gene, which is essential for O-glycan elongation [O-glycans in MKN45 SC cells and AGS SC cells promoted invasive features. The results of the transcriptomic analysis exhibited that COSMC knockout in MKN45 SC cells and AGS SC cells resulted in a significant alteration in the transcription profile in comparison with the respective parental cells, especially the most upregulated SRPX2 gene, suggesting that SRPX2 may promote cancer invasion through regulating cell glycosylation [Glycosylation is the most ubiquitous molecular modification, which regulates many biological functions . Compareruncated , 45. Abnruncated . MKN45 Songation , 48. Fresylation .Cancer resistance is still the greatest challenge in improving the outcomes for cancer patients . AccumulResearchers have found SRPX2 may yield several clinically relevant insights in patients with cancers. Li and coworkers demonstrated that patients with elevated SRPX2 expression are related to shorter disease-free survival (DFS) and overall survival (OS) than patients with low SRPX2 expression by detecting 200 tissue microarray specimens from patients  with curative pancreatectomy for pancreatic ductal adenocarcinoma . Furthervia the TGFβR1/SMAD3/SRPX2/AP1/SMAD7 pathway in vitro and in vivo. Thus, these findings indicated that SRPX2 might be a potential therapeutic target for treatment of highly metastatic cancers, cancer resistance, and inflammation. Inflammatory signaling pathways are closely involved in tumorigenesis, therapy resistance, and metastasis. A better understanding of the roles of SRPX2 in inflammation will be helpful to treat cancer-related inflammation to slow the progression of cancer. Additionally, it is a pity that there is little connection between SRPX2 and drug development at current. Therefore, in-depth understanding of structure and functions of SRPX2 will provide the theoretical foundations to drug design.Since its discovery in 1999, SRPX2 is demonstrated to associate with the progression of various types of cancers. Research interest in cancer mainly focused on cancer metastasis, invasion, migration, and drug resistance through regulating the FAK signaling pathway, EMT, angiogenesis, and glycosylation . Moreove


Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disease in the world. Sustained hepatic inflammation is a key driver of the transition from simple fatty liver to nonalcoholic steatohepatitis (NASH), the more aggressive form of NAFLD. Hepatic inflammation is orchestrated by chemokines, a family of chemoattractant cytokines that are produced by hepatocytes, Kupffer cells (liver resident macrophages), hepatic stellate cells, endothelial cells, and vascular smooth muscle cells. Over the last three decades, accumulating evidence from both clinical and experimental investigations demonstrated that chemokines and their receptors are increased in the livers of NAFLD patients and that CC chemokine ligand (CCL) 2 and CCL5 in particular play a pivotal role in inducing insulin resistance, steatosis, inflammation, and fibrosis in liver disease. Cenicriviroc (CVC), a dual antagonist of these chemokines’ receptors, CCR2 and CCR5, has been tested in clinical trials in patients with NASH-associated liver fibrosis. Additionally, recent studies revealed that other chemokines, such as CCL3, CCL25, CX3C chemokine ligand 1 (CX3CL1), CXC chemokine ligand 1 (CXCL1), and CXCL16, can also contribute to the pathogenesis of NAFLD. Here, we review recent updates on the roles of chemokines in the development of NAFLD and their blockade as a potential therapeutic approach. N-terminals that are key to forming their 3-dimensional shape [Chemokines  are a large family of chemotactic cytokines that involve immune and inflammatory responses through the chemoattraction and activation of leukocytes . These sal shape . To dateal shape .Chemokine receptors are a group of ~20 typical G protein-coupled seven-transmembrane proteins and are expressed in various leucocytes and immune cells. The directed migration of specific chemokine receptor-expressing cells allows for their recruitment along a chemokine concentration gradient . Upon liRegarding the pathogenesis of NAFLD, relevant chemokines and their receptors have been well summarized by excellent reviews ,12. PartIn this review, we will highlight recent updates on the roles of chemokines, including CCL3, CCL25, CXCL1, CXCL16, and CX3CL1 , in the CCL3 (known as macrophage inflammatory protein-1α) is expressed in macrophages and secreted to recruit macrophages themselves, various leukocyte subtypes, and T cells to inflamed sites ,24. VariThe progression of some inflammatory diseases, including atherosclerosis , rheumatWe further investigated the role of CCL3 in the pathogenesis of NAFLD in mice fed a high-cholesterol and high-fat (CL) diet, a dietary model of NASH . We foun+CD4+ tissue-infiltrating T cells in both Crohn’s disease and a murine model of inflammatory bowel disease [+ macrophages play a pathogenic role in a murine acute hepatitis model and humans [+ monocytes than healthy volunteers [+ macrophages accumulate in the liver, which produces high levels of TNF-α and promotes the Th1 differentiation of naive CD4+ T cells, thereby contributing to acute liver inflammation [+/LYVE1+ sinusoidal endothelial cells, whereas CCR9 was expressed in CD68+ macrophages and GFAP+/α-SMA+ HSCs in the livers of patients with NASH, and the numbers of these CCR9+ cells were significantly lower in the control samples. (3) CCR9-deficient mice showed alleviated diet-induced steatohepatitis associated with the decrease in the amount of CD11b+ inflammatory macrophage accumulation in the liver. (4) Consistent with human NASH, CCR9 was also expressed on HSCs in NASH mice and CCR9-deficient HSCs show fewer fibrogenic phenotypes. Finally (5) A CCR9 antagonist, vercirnon (CCX282-B) ameliorated steatohepatitis and the development of diethylnitrosamine-induced hepatocellular carcinoma in a high-fat diet-fed mice. These results indicate a therapeutic potential of CCR9 blockade in NAFLD.The chemokine CCL25 is selectively and constitutively expressed in the thymus and small intestine. CCR9, the sole functional receptor of CCL25 , is expr disease ,38,39. Wd humans . Periphelunteers . Similarammation . Additioammation : (1)SeruCXCL1 is one of the major chemoattractants for neutrophils . After b+ T cells [+ NKT cells patrol liver sinusoids and provide the intravascular immune surveillance of pathogens [+ cells to cholangiocytes and hepatocytes by triggering the conformational activation of β1 integrins and the binding to vascular cell adhesion molecule-1 (VCAM-1), thereby promoting liver inflammation [In conjunction with CD4, CXCR6 can serve as a co-receptor for the entry of human and most simian immunodeficiency viruses (human immunodeficiency virus type I and simian immunodeficiency virus) . Similar T cells . In the athogens . CXCL16,athogens . CXCL16 ammation . Regardiammation . Additioammation , suggest+ T cells. The auto-aggression of CD8+ T cells may be involved in the development of hepatocellular carcinoma from NASH. CXCR6+ CD8+ T cells accumulate in the livers of a preclinical mouse model of NASH (mice fed a choline-deficient and HFD) and of patients with NASH [In the liver, CXCR6 is also expressed in CD8ith NASH . The T cith NASH .N-terminal domain of CX3CL1, containing a CX3C motif, can be cleaved by ADAM Metallopeptidase Domain 10 (ADAM10) [CX3CL1, also known as fractalkine, a membrane-anchored chemokine, is expressed on epithelial cells, dendritic cells, and neurons and could be induced by inflammatory cytokines, such as TNF-α and IFN-γ ,59,60,61(ADAM10)  and ADAM(ADAM10) , yieldin(ADAM10) .gfpgfp/) showed less moDC recruitment into the liver associated with incomplete maturation of monocytes into moDCs [−/−) mice exposed to CCl4 exhibited increased inflammatory cell recruitment into the liver and pro-inflammatory cytokine/chemokine production, including TNF-α, IL-1β, CCL2, and CCL5, but decreased expression of anti-inflammatory IL-10 and arginase-1 in Kupffer cells, resulting in enhanced HSC activation and subsequent liver fibrosis [−/− mice were more prone to HFD-induced obesity, insulin resistance, and hepatic steatosis and inflammation compared with wild-type control mice [® vector  alleviated insulin resistance and inflammation in the liver and eWAT of obese mice [Similar to the other chemokines, both animal and clinical studies have demonstrated that CX3CL1-CX3CR1 signaling is enhanced in various inflammatory diseases, such as rheumatoid arthritis , atherosto moDCs . Additioto moDCs . These rfibrosis . Additiorol mice . We alsoese mice . CollectSince CCR2 and CCR5 play an important role in the infiltration of myeloid cells and the activation of HSCs, CVC, a once-daily, orally available CCR2/CCR5 dual antagonist, has been expected to improve NASH by suppressing both inflammation and fibrosis, as shown in animal models of steatohepatitis ,78. In tCitrus unshiu (Satsuma mandarin orange) and Solanum lycopersicum (tomato), respectively, are relatively abundant in human blood [Many epidemiological studies have demonstrated that the development of NAFLD is closely linked to lifestyle factors  . A nutrian blood ,86,87 anan blood ,89,90. Tan blood ,89.Sulforaphane, an isothiocyanate derived from cruciferous vegetables, such as broccoli, is a potent inducer of nuclear factor (erythroid-derived 2)–like 2 (Nrf2), a master transcription factor that regulates oxidative stress responses ,91. In aAccumulating evidence from in vitro and in vivo studies reveals that some chemokine-chemokine receptor axes play a central role in liver inflammation during the development of NAFLD. However, as the results from the AURORA study demonstrate, therapeutic applications for targeting chemokines and chemokine receptors to resolve steatohepatitis and fibrosis are still challenging. Further basic and clinical research is essential for better understanding the molecular mechanisms by which the chemokine system mediates hepatic and adipose inflammation as well as their interaction in the progression of NAFLD. To improve the response rates among patients with NAFLD, combination approaches including lifestyle interventions that are personally tailored to the patient’s disease drivers, such as obesity and type 2 diabetes, are required.


Interestingly, our results also demonstrate that cadherin-11 is required to preserve gp130 levels for IL6 family signaling. At the same time, however, activated Src527F downregulates cadherin-11, in a quantitative manner. As a result, Src527F expression to intermediate levels allows sufficient cadherin-11, hence gp130 levels for Stat3 activation, as expected. However, expressed to high levels, Src527F eliminates cadherin-11, hence gp130 signaling, thus abolishing Stat3-ptyr705 stimulation. Taken together, these data establish for the first time a loop between Src, cadherin-11, gp130, and Stat3 activation. This fine balance between Src527F and cadherin-11 levels which is required for Stat3 activation and cellular survival could have significant therapeutic implications.We previously demonstrated that engagement of cadherins, cell to cell adhesion molecules, triggers a dramatic increase in levels and activity of the Rac/Cdc42 small GTPases, which is followed by secretion of IL6 family cytokines and activation of their common receptor, gp130, in an autocrine manner. This results in phosphorylation of the Signal Transducer and Activator of Transcription-3 (Stat3) on tyrosine-705, which then dimerizes, migrates to the nucleus, and activates transcription of genes involved in cell division and survival. In the present report we demonstrate that, in mouse Balb/c3T3 fibroblasts, mutationally activated Src This527F. These cells are not transformed so that, unlike tumor cells, the relationship between Src, cadherin-11, and Stat3 can be examined in the absence of confounding pathways. Our results reveal the importance of a fine balance between Src527F and Cad11 levels for Stat3 activation; expressed to high levels, Src527F eliminates Cad11, which, in turn, is required for gp130 signaling and Stat3 activation. Expressed to intermediate levels, on the other hand, Src527F allows sufficient residual levels of Cad11 and gp130 for Stat3 activation. Taken together, these data establish a loop between Src, Cad11, gp130, and Stat3 activation, a finding which could have significant therapeutic implications.The Src family of non-receptor tyrosine kinases is often hyperactive in a variety of cancers , and act2 incubator. Balb/c3T3 cells and derivatives required supplementation with 10% fetal bovine serum , but the rat-F111 cells and derivatives required 5% calf bovine serum. It was especially important to ensure even distribution of the cells during plating, by passing them at subconfluence and pipetting vigorously with a 9” Pasteur pipette. To reduce the variability that might be caused by nutrient depletion in post-confluent cultures, the medium was changed every 24 h. Cell confluence was estimated visually and quantitated by imaging analysis of live cells under phase contrast using a Leitz Diaplan microscope and the MCID-elite software .The Balb/c3T3 cell line (ATCC) has been described . The ratAll lines were treated with plasmocin  periodically and tested for mycoplasma contamination by DAPI staining or by PCR, as described in .527F-hygro plasmid which encodes the constitutively active chicken Src mutant and a hygromycin resistance gene was a gift from Dr. Andrew Craig, Queen’s University and propagated by transfection in Phoenix ecotropic retroviral packaging cells using the PolyJet in vitro DNA transfection reagent . Following selection, clones picked were further subcloned at least twice more to eliminate revertants and propagated at subconfluence. A number of vials of each were frozen and used for a maximum of two weeks after thawing. This was especially important for high-Src527F-expressing cells.The pWZL-SrcBalb-shCad11 and Balb-shRac cells have been described , 20. CadpY416 , pan-Src , Cadherin-11 , Stat3-ptyr705  cleaved PARP , Rac (Millipore #05-389), b-actin  and secondary antibodies according to the manufacturer’s protocols. Bands were visualized using the Clarity Western ECL substrate . Images were developed by exposing the membranes to film or by using the Azure 300 Digital imager . Photo-shop or Corel Draw software was used for the organization of non-adjusted, original images. Quantitation was achieved by image analysis using ImageJ (U.S. NIH). In all cases, band intensities were normalized to β-actin or α-tubulin levels of the same samples. Rac activation assays were performed using GST-PAK pull-down assays with the Rac activation kit from Cytoskeleton , using beads coated with glutathione-S-transferase (GST) fused to the binding domain of p21-activated kinase (PAK), followed by probing for Rac by Western blotting, as previously described [Following electrophoresis and transfer, membranes were cut into strips based on the positions of pre-stained markers and probed for SrcExperiments were repeated at least twice and with a number of lines expressing different Src levels. Stat3-ptyr705 levels were examined at different densities spanning 100% confluence, as described in Results.Cells were fixed with 4% paraformaldehyde and assayed for apoptosis by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) using fluorescein-coupled nucleotides, according to the manufacturer’s instructions . Cells with apoptotic nuclei were visualized and photographed under fluorescence and phase contrast illumination.527F was stably expressed in mouse Balb/c3T3 fibroblasts which possess high Cad11 levels [pY416 (which correlates with Src activity) by Western blotting , Src-med cells had approximately 42% the Cad11 protein levels of the parental Balb/c3T3 cells (lane 3 vs 1), while Src-low cells had approximately 75% the Cad11 levels of the Balb/c3T3 (lane 2 vs 1). A number of additional clones expressing different Src527F levels gave similar results  is a potent Stat3 activator. To examine this apparent paradox, we assessed the effect of different levels of Src527F upon the Cad11/Stat3 axis. Since cadherin engagement, as occurs with a density of cultured cells, causes a dramatic increase in Stat3 activity [[med, and Src-high cells were plated in plastic culture dishes and, when 50% confluent and over several days thereafter  to Src-med cells , in agreement with previous findings [527F expression (line Src-high) rather than increasing, dramatically decreased Stat3-ptyr705 to undetectable levels (lanes 11–15). A number of additional clones expressing graded Src527F levels gave similar results , indicating that although Src527F can activate Stat3 in the parental Balb/c3T3 when expressed to medium levels   and fibr527F, we explored whether Rac might actually be involved in Stat3 activation by Src527F as well. At first, the effect of Src upon Rac protein levels was examined. Balb/c3T3, Src-med, and Src-high cells were plated at densities of 100% confluence or 100% plus one day and Rac levels assessed by Western blotting. As shown in Fig. 527F expression triggered an increase in Rac protein levels, which was most prominent at high Src527F levels . Activated Rac-GTP mirrored the levels of Rac protein (lanes 7–9). Taken together, these data indicate that Src527F expression is able to increase Rac protein levels and activity. Moreover, this increase must be independent of any effects of Cad11, since in Src-high cells Cad11 is undetectable while Rac is the highest.Since, as shown above, Cad11 is required for Stat3 activation by Src527F. Rac was downregulated through the expression of shRac with a retroviral vector . As shown in Fig. 527F expression in shRac cells caused only a small (12%) increase in Stat3-ptyr705 (lane 4), compared to Src527F expression in the parental Balb/c3T3 (lane 2), revealing a Rac requirement for Stat3 activation by Src527F. It is possible that Src may be using Cdc42 as an alternate branch to transmit the signal to gp130/Stat3, which could explain the background level of Stat3-ptyr705 (12%) in shRac-Src-med cells (lane 4). Taken together, these findings indicate that Src527F increases Rac levels in Balb/c3T3 fibroblasts, by a mechanism that is independent of Cad11. In addition, Rac is, in fact, required for full Stat3, ptyr705 phosphorylation following Src527F expression, consistent with previous reports [We next examined the Rac requirement for Stat3 activation by Src reports .527F expression, we assessed the secretion of cytokines by cells expressing different Src527F levels. Balb/c3T3, Src-med, or Src-high cells were grown to high densities and conditioned medium collected 24 h later. This medium was added to Balb/c3T3 cells grown to a low density (50% of confluence) and Stat3-ptyr705 levels were assessed 30 min later by Western blotting . These results indicate that Src-med cells may secrete high amounts of cytokines, which, acting in an autocrine manner, may induce the high Stat3-ptyr705 levels observed in Src-med cells, as shown in Fig. high cells also increased Stat3-ptyr705 in sparsely growing Balb/c3T3 cells, pointing to the secretion of high amounts of cytokines (lane 4), despite the fact that their own Stat3-ptyr705 levels were undetectable , while high Src expression levels eliminated gp130 altogether (lanes 9–12), concomitant with the elimination of Stat3-ptyr705. Interestingly, Stat3-ptyr705 levels were higher in Src-med cells (lanes 5–8) than Balb/c3T3, despite their lower gp130 levels, presumably due to higher Rac and IL6 family cytokines secreted by Src-med than the parental Balb/c3T3. However, high Src527F levels that eliminated Cad11 and gp130, eliminated Stat3-ptyr705 as well (lanes 9–12). Levels of Jak1-p1022/1023 mirrored levels of gp130 and Stat3  above the DMSO controls. Interestingly, however, treatment of Src-high cells with Dasatinib caused a prominent gp130 increase, which translated into a robust increase in Stat3-ptyr705 (lanes 8 and 10 vs 7 and 9). These results confirm that expressed to high levels, Src actually reduces gp130 and Stat3 activity. Thus, the exact level of Src expression in a given cancer could offer a potential explanation for the inconsistent results of Dasatinib or other Src inhibitors in clinical trials; treatment of high-Src cancers would actually increase Stat3 activity, leading to metastasis.Despite the fact that Src has been found to be hyperactive in a number of cancers, Src inhibitors such as Dasatinib were found to be largely ineffective in clinical trials , 29. It bcl-xL, mcl-1, and survivin [527F, besides Stat3, is known to activate the E2F transcription factor family, which actually promotes apoptosis, through the Ras/Raf/Erk pathway [527F expression upon apoptosis vs survival, which could be different depending upon Src activity levels. To answer this question, cells were grown to 1 day post-confluence and apoptosis examined by TUNEL staining and analysis of poly (ADP-ribose) polymerase (PARP) cleavage by Western blotting. As shown in Fig. med cells displayed a very low level of apoptosis compared to the parental Balb/c3T3 , rather than resulting in even higher Stat3-ptyr705 levels, was found for the first time to eliminate Stat3-ptyr705 entirely. The fact that Cad11 downregulation indeed prevented Stat3, ptyr705 phosphorylation by Src527F  increases Rac protein levels as well. Interestingly, we previously demonstrated that Rac, upregulated either through cadherin engagement [V12) increases transcription of IL6 family cytokines which stimulate the gp130 receptor in an autocrine manner, leading to an increase in activity of Jak and Stat3 [high cells , which The observation that in cultured cells, and potentially in vivo, high Src levels downregulate Cad11, which, in turn, may be required for metastasis, especially to the bone, may signify that actual metastatic tumors cannot possibly express such high Src levels as to eliminate Cad11. Apparently, a fine balance between Src and Cad11 levels is required, as to allow both transformation (through Src) and metastasis (through Cad11), for the formation of a metastatic tumor to occur. As a result, pharmacological inhibition of Src would increase Cad11/gp130/Stat3 levels in tumors with high Src activity, hence increase, rather than decrease, the metastatic ability of the tumor, leading to a deterioration in clinical outcome. The poor performance of Src inhibitors including Dasatinib in clinical trials is consistent with these findings –59.Our results also stress for the first time the importance of Cad11, Rac, and IL6/gp130 signaling in Src-triggered induction of Stat3, tyr705 phosphorylation, and activation. As such, Cad11 and likely other cadherins are emerging as not only powerful Stat3 activators, but also as critical mediators of Stat3 signaling from Src and possibly other oncogenes, despite the fact that, at the same time, Src downregulates cadherins. Conversely, downregulation of Stat3-ptyr705 by caveolin-1, the main protein of caveolae, has been shown to be mediated through the downregulation of Cad11 . FurtherSupplementary Data


With the increasing incidence of metabolic diseases year by year and their impact on the incidence of cardiovascular diseases, metabolic diseases have attracted great attention as a major health care problem, but there is still no effective treatment. Oxidative stress and inflammation are the main mechanisms leading to metabolic diseases. T cells are involved in the inflammatory response, which can also regulate the development of metabolic diseases, CD4+ T cells and CD8+ T cells are mainly responsible for the role. Th1 and Th17 differentiated from CD4+ T promote inflammation, while Th2 and Treg inhibit inflammation. CD8+ T cells also contribute to inflammation. The severity and duration of inflammatory reactions can also lead to different degrees of progression of metabolic diseases. Moreover, mTOR, PI3K-Akt, and AMPK signaling pathways play unique roles in the regulation of T cells, which provide a new direction for the treatment of metabolic diseases in the future. In this review, we will elaborate on the role of T cells in regulating inflammation in various metabolic diseases, the signaling pathways that regulate T cells in metabolic diseases, and the latest research progress. Metabolic disease is a kind of multi-system abnormal disease which is manifested by diseases or disorders that disrupt normal metabolism, including hyperglycemia, dyslipidemia, hypertension, obesity, and insulin resistance, and leads to a dramatic increase in the occurrence of cardiovascular diseases such as acute myocardial infarction and stroke . CompareT cells play an indispensable role in inflammation and are directly involved in the development of metabolic diseases, for example: (1) During chronic inflammation, the balance of Th17/Treg is closely related to metabolic diseases . (2) T cT lymphocytes originate from bone marrow, mature in the thymus, then differentiate into two major T cell lineages with different immune functions. A small fraction belongs to γδ-lineages, and migrate from the thymus to the epidermis, mucosa, and intestine, acting as the natural surveillance cells of the immune system, but most T cells belong to αβ-lineage . ImmaturIn the early phase of inflammation, cytokines such as IL-6 and tumor necrosis factor-α (TNF-α) are produced by T cells, as inflammatory stimuli, which can be recognized by pattern-recognition receptors. After that, inflammatory pathways including nuclear factor kappa-B (NF-κB) pathway, mitogen-activated protein kinase (MAPK) pathway, and Janus kinase signal transducer and activator of transcription (JAK-STAT) pathway are activated. In addition, the NF-κB pathway can regulate the biological processes of T cell-derived pro-inflammation factors such as IL-6, interferon-γ (IFN-γ), and granulocyte macrophage-colony stimulating factor (GM-CSF). However, acute inflammation is mainly regulated by innate immunity, and the recruitment of effector cells to infectious sites is the master ingredient of innate immunity. The γδ T cells could directly kill infectious cells through releasing cytotoxic granules and granulysin. Meanwhile, they can indirectly eliminate microbes by secreting pro-inflammation cytokines . Most imAcute inflammatory state may switch to chronic state when immune cells are unable to eliminate pathogens . Antigenhttps://biorender.com, accessed on 7 September 2022.).During acute inflammation, T cells are activated after T lymphocytes recognize the antigen. Activated T cells secrete cytokines  which can participate in recruiting macrophages and produce IFN-γ to activate macrophages. The activated macrophages stimulate T lymphocytes by antigen presentation and different cytokines at the same time. During chronic inflammation, CD8+ T cells express inhibitory receptors such as PD-1 and CTLA-4, then exhausted, and the effector function and proliferative capacity reduce. TNF: tumor necrosis factor; IL-17: interleukin-17; PD-1: programmed cell death protein 1; CTLA-4: cytotoxic T lymphocyte-associated antigen-4. The figure was constructed with BioRender  pathways, which is attributed to the suppression of inflammatory signals ,42. MeanMoreover, inflammation also promotes tissue remodeling. These responses involve stimulating angiogenesis to combat hypoxia, developing insulin resistance to protect adipocytes from lipids overaccumulation , and exphttps://biorender.com, accessed on 7 September 2022).Pro-inflammatory cytokines promote the recruitment the macrophage and secretion of TNF to act on metabolic diseases including insulin resistance, β cell dysfunction, impaired glucose tolerance, and tissue remodeling. TNF: tumor necrosis factor. The figure was constructed with BioRender  cells ,53. Th1 IFN-γ is produced by Th1 cells. Studies showed that the level of IFN-γ is higher in high-fat diabetes (HFD)-induced obese mice and obese populations . FurtherIL-17 is mainly secreted by Th17 cells, which suppresses glucose metabolism and adipogenesis and postpones obesity progression in mice and humans . A studyTregs exhibit an anti-inflammatory effect via IL-10 , which hhttps://biorender.com, accessed on 7 September 2022).Naïve CD4 T cells differentiate into different subsets in various circumstances, including Th1, Th2, Th17, and Treg cells. Generally, Th1 and Th17 cells secrete pro-inflammatory cytokines, and Th2 and Treg cells produce anti-inflammatory cytokines. The figure was constructed with BioRender (CD8+ T cells differentiate into cytotoxic T lymphocyte (CTL) after being stimulated by antigens. CTL kills target cells by secreting cytotoxic substances such as perforin and granzyme, and induces apoptosis through the Fas/FasL pathway . It was CD8+ T cell infiltration increase in both obese humans and mice with nonalcoholic fatty liver disease (NAFLD) . In mousNishimura et al. discovered that the accumulation of CD8+ T cells results in insulin resistance and chronic inflammation in mice . Chronichttps://biorender.com, accessed on 7 September 2022).CD8+ T cells play a role in pro-inflammatory activities. CD8+ T cells infiltration promotes the activation and recruitment of macrophages. Moreover, CD8+ T cells infiltration in liver tissue is beneficial to the conversion from NASH to hepatocarcinoma. Moreover, cumulated exhausted CD8+ T cells suppress the insulin sensitivity which leads to insulin resistance and impaired glucose tolerance. NASH: non-alcoholic steatohepatitis. The figure was constructed with BioRender  . This mamTOR is important in metabolic diseases. An increased proportion of Treg / Th17 can reduce AT inflammation and insulin resistance caused by obesity. In terms of NAFLD, the inflammation of liver can be reduced by alleviating liver inflammation and fibrosis, thereby inhibiting the progress of T2D and promoting the islets’ health, finally preventing inflammation caused by NAFLD.The PI3K/Akt signaling pathway is associated with proliferation, differentiation, and apoptosis, which is crucial in organismal growth and critical cellular processes by mediating growth factors . ParticuPI3Ks are a family of lipid kinases . PI3Ks cThe PI3K signal pathway is essential for native CD4 T cells to differentiate into the main Th cell subsets, Th1, Th2, Th17, and TFH ,121,122.PI3K plays an important role in Th1 and Th2 differentiation through complex mechanisms. mTOR is a crucial signaling protein to connect PI3K activation and Th differentiation. In mouse T cells with mTOR knockout, clonal proliferation of mouse T cells was reduced and differentiation into Th1, Th2, or Th17 subsets was completely inhibited . ConversRecent studies suggested that the development of nTregs is also regulated by PI3K-Akt- Forkhead Box O (FOXO). Moreover, it has been reported that the combined deletion of Foxo1 and Foxo3 can suppress the development of FoxP3+ nTregs in the thymus ,127. WheAdditionally, it is reported that restriction of PI3K/Akt/TOR activity promotes CD4 T cells to convert to iTregs . Other wFurthermore, activated CD8+ T cells require PI3K/Akt signaling to initiate the transcription function of CTL. Akt activity and FOXO inactivation are associated with the expression of cytotoxicity effectors, including perforin, granzymes, and IFN-γ . AKT actEvidence has shown that PD-1-deficient Treg cells can inhibit PI3K/Akt signaling, thereby enhancing Treg immunosuppressive ability, and in mice that only have a PD-1 deficient in Tregs, their diabetes has improved . Togethe2+ respectively, while the tumor suppressor gene FLCN inhibits activation of AMPK, phosphorylation of TSC2 (activation) and RAPTOR (inhibition) downstream of AMPK inhibit mTOR activity, which in turn reduces mRNA translation [AMPK is an AMP-activated protein kinase, which is a key molecule in the regulation of bioenergy metabolism, and plays an indispensable role in T cell metabolism and function . In the nslation . The catnslation .A function of AMPK towards T cells is regulating fuel usage, as activated T cells possess glucose-sensitive metabolic checkpoints controlled by the energy sensor AMPK, which regulates messenger RNA (mRNA) translation as well as mitochondrial metabolism. Blagih et al. noted that the capacity of GU and the function of mitochondria were reduced in AMPKα1-deficient T cells, which leads to decreased effector T cell responses to pathogens . Pearce Also, research carried by Nyambuya showed that metformin has anti-inflammatory effects by activating the AMPK signaling pathway and inhibiting the mTOR signaling pathway, thereby slowing down the progression of T2D and T2D-related cardiovascular complications .Metabolic diseases are common worldwide; popular examples include obesity and NAFLD. Inflammation and oxidative stress are the main pathogenesis of metabolic diseases, and chronic or excessive inflammation can affect their progression, and T cells play an indispensable role in inflammation. In this review, we focused on the role of T cells in regulating inflammation in different metabolic diseases and the role of signaling pathways in regulating T cells, as well as the latest research progress and future direction. In metabolic diseases, such as obesity, NAFLD andT2D, CD4+ T cell differentiated cells, such as Th1, Th2, Th17, and Tregs, all play potential roles in disease regulation or progression. However, there are still several mechanisms worth studying. The functions of Th2 cells in metabolic diseases are not well understood , and theLastly, translating our knowledge of these signaling pathways in T cells into therapeutic modalities remains a challenge in the future. Future studies on the regulation of T cells by numbers of signaling pathways will facilitate the development of targeted immunotherapies for multiple metabolic diseases.


NOL7 in cancer initiation and development, a systematic pancancer analysis of NOL7 is lacking. Herein, we present a comprehensive study of NOL7 which aimed to explore its potential role and detailed mechanisms across 33 human tumors based on The Cancer Genome Atlas (TCGA) and Clinical Proteomic Tumor Analysis Consortium (CATPAC) databases. As a result, both gene and protein levels of NOL7 were found to be increased in various tumor tissues, including breast invasive carcinoma (BRCA), colon adenocarcinoma (COAD), hepatocellular carcinoma (LIHC), lung adenocarcinoma (LUAD), and head and neck squamous cell carcinoma (HNSC) as compared with corresponding normal tissues. Meanwhile, dysregulated NOL7 expression was found to be closely related to pathological stage and prognosis in several cancers, including LIHC, ovarian serous cystadenocarcinoma (OV), and bladder urothelial carcinoma (BLCA). The DNA methylation level of NOL7 was found to be decreased in most cancers and to be negatively associated with NOL7 expression. Furthermore, NOL7 expression was determined to be significantly associated with levels of infiltrating cells and immune checkpoint genes, including HMGB1. Analysis of NOL7-related genes revealed that RNA metabolism pathways, including “ribosome biogenesis”, “spliceosome”, and “RNA transport”, were mainly involved in the functional mechanism of NOL7 in human cancers. In summary, this pancancer study characterized the relationship between NOL7 expression and clinicopathologic features in multiple cancer types and further showed its potential regulatory network in human cancers. It represents a systemic analysis for further functional and therapeutic studies of NOL7 and highlights its predictive value with respect to the carcinogenesis and prognosis of various cancers, especially LIHC.Despite growing evidence suggesting the critical function of  Targeted therapy has become mainstream and one of the most effective strategies for cancer treatment . HoweverNOL7 gene is located at 6p23, a chromosomal region frequently lost in several malignancies, including cervical cancer [NOL7 serves as a tumor suppressor gene in cervical cancer [NOL7 were observed in cervical cancer, and it was found to be downregulated with disease progression. Meanwhile, NOL7 can predict cancer recurrence and survival in cervical cancer patients. Researchers found that tumor-associated mutations and SNP variations harbored in the NOL7 gene potentially contribute to the loss of NOL7 in cancer cells [NOL7 expression [NOL7 was positively regulated by the RB tumor suppressor in cervical cancer and that it further functioned as an mRNA-binding protein to modulate TSP-1 expression by enhancing the post-transcriptional stability of its mRNA, thus inducing an anti-angiogenic and tumor-suppressive phenotype [NOL7 gene encodes nucleolar protein 7, which has three alternatively spliced forms, each isoform differently localized within the cell. NOL7-SP1 (29 kD) is predominantly localized in the cell nucleolus, with some nucleoplasmic distribution; NOL7-SP2 (16 kD) is nucleoplasmic and has a moderate nucleolar distribution; while NOL7-SP3 (12 kD) is mostly distributed in the nucleoplasm, with some cytoplasmic distribution. A strong nuclear localization signal within the C-terminus of NOL7-SP1 contributes to its translocation into the cell nucleus. Nucleolus-localized NOL7 protein is typically involved in maintaining internal nucleolar structure and cell growth, whereas nucleoplasm-localized NOL7 protein exerts anti-angiogenic effects [NOL7 gene resides, frequently undergoes heterozygous loss in cervical cancer, and this region is commonly associated with the amplification of melanoma [NOL7 was upregulated in melanoma development and exhibited tumor-promoting activity [NOL7 suggested that its investigation might enable mechanistic insights into the potential role of NOL7 in malignancy. However, a systematic assessment of NOL7 in human cancer remains unavailable. The human l cancer . It was l cancer . Both lopression . MW Linghenotype ,9. The N effects ,11. It iNOL7-SP3  kD is moactivity . The mulNOL7 in cancer, we first combined TCGA project, GTEx, and CPTAC datasets to conduct a pancancer analysis of NOL7 in various human cancers. This comprehensive study aimed to clarify the potential function and regulatory mechanisms of NOL7 in the pathogenesis or clinical prognosis of multiple cancer types and further confirm its promising role as a diagnostic biomarker and therapeutic target for cancer treatment.Given the critical role of NOL7 gene is localized at the 6p23 chromosomal region . We downloaded data on the subcellular localization and tissue localization of the NOL7 protein from the HPA database and found that the NOL7 protein had the highest expression in the kidneys, followed by the pancreas , COAD, esophageal carcinoma (ESCA), head and neck squamous cell carcinoma (HNSC), LIHC, LUAD, lung squamous cell carcinoma (LUSC), prostate adenocarcinoma (PRAD), and stomach adenocarcinoma (STAD) . Conversely, decreased NOL7 expression was detected in kidney chromophobe (KICH), kidney renal clear cell carcinoma (KIRC), kidney renal papillary cell carcinoma (KIRP), and thyroid carcinoma (THCA) tissues  as compared with normal tissues. We also incorporated the GTEx datasets to supplement the number of normal tissues included as controls in the TCGA project. As shown in NOL7 levels were significantly higher in thymoma (THYM), CHOL, DLBC, LIHC, and pancreatic adenocarcinoma (PAAD) tissues as compared with corresponding normal tissues, while they were markedly lower in acute myeloid leukemia (LAML) tissues as compared with normal tissues .Moreover, a gene expression landscape of NOL7 protein levels between tumor tissues and corresponding normal tissues. The protein levels of NOL7 were found to be notably increased in breast cancer, ovarian cancer, colon cancer, hepatocellular carcinoma, KIRC, adrenocortical carcinoma (RCC), LUAD, glioblastoma multiforme (GBM), and HNSC tissues  but decreased in PAAD tissues (p < 0.05) as compared with relevant healthy tissues , KIRP (p < 0.001), LIHC (p < 0.01), sarcoma (SARC) (p < 0.001), and UCEC (p < 0.05). Among all tumor types, KIRP showed the greatest significance. In contrast, reduced NOL7 expression was related to favorable OS in bladder carcinoma (p < 0.05), KIRC (p < 0.01), ovarian cancer (p < 0.001), and READ (p < 0.001). p < 0.001) and ovarian cancer (p < 0.01). In addition, increased NOL7 expression corresponded with poor distant metastasis-free survival (DMFS) (p < 0.05) and RFS (p < 0.001) in breast cancer but was related to better OS (p < 0.001), first progression (FP) (p < 0.001), and post-progression survival (PPS) (p < 0.001) in gastric cancer. Additionally, increased NOL7 expression was remarkably related to poor PFS (p < 0.001) and better PPS (p < 0.01) in ovarian cancer and to better FP (p < 0.001) in lung cancer ; the indications were opposite for other cancers, including ACC, ESCA, KICH, SARC, and LIHC . However, augmented NOL7 expression predicted a good prognosis in terms of OS in both KIRC and OV . Taken together, these results demonstrated that NOL7 expression could predict patient survival in several cancer types, especially ACC, KICH, KIRC, LIHC, LUAD, and OV. However, the specific relationship between NOL7 expression level and prognosis in cancer patients depends on the tumor type.According to the results from GEPIA2, we observed that dysregulated NOL7 expression significantly affected OS in seven types of cancer. For KIRC and OV, a favorable prognosis was presented when NOL7 expression was increased , KIRP, and ESCA  as compared with normal tissues  for NOL7. Amplification type variation accounted for a significant portion of the alterations in OV, DLBC, CHOL, uveal melanoma (UVM), UCS, and HNSC b. The inTumor-infiltrating immune cells play a key role in the progression and invasion of cancer . Given tWe further explored the correlations between NOL7 expression and TMB and MSI. A positive correlation between NOL7 expression and MIS was observed in GBM, brain lower-grade glioma (LGG), stomach and esophageal carcinoma (STES), SARC, STAD, UCEC, HNSC, KIRC, LUSC, and UCS. In addition, NOL7 expression was positively correlated with TMB in LUAD, LAML, BRCA, ESCA, STES, SARC, STAD, LUSC, THYM, LIHC, PAAD, SKCM, UCS, BLCA, and ACC .p-values < 0.01) , EEF1E1 (R = 0.61), SSB (R = 0.57), DHX9 (R = 0.53), and WRNIP1 (R = 0.6)  a, above. < 0.01) a, below. < 0.01) c. NOL7 on tumor pathogenesis  compared with normal tissues. Integration of these data with CPTAC datasets revealed that both gene and protein expression levels of NOL7 were significantly enhanced in breast cancer, colon cancer, hepatocellular carcinoma, LUAD, and HNSC. Consistently, both gene and protein expression of NOL7 were upregulated, along with disease progression in various cancers, including colon cancer, LUAD, and HNSC  of the and HNSC . These rncreased . It is wC and OV . Based oNOL7 through loss of heterozygosity and decreased mRNA and protein expression has been observed in cervical cancer [NOL7 in cervical cancer [NOL7 resides, is frequently amplified in several cancers, including melanoma, and therefore may exert disease-promoting effects in these cancers [NOL7 expression was found in metastatic sites as compared with primary sites in SKCM, suggesting the promotion of melanoma development. These findings were consistent with our previous conclusion [NOL7 promotes carcinogenesis in hepatocellular carcinoma awaits verification [Previous researchers have confirmed that consistent loss of  cancers . In thisnclusion , whereasfication . NOL7 protein is required to maintain internal nucleolar structure and cell proliferation [NOL7-encoding protein is an essential molecule for cytogenetic events and that loss of NOL7 leads to the fatal destruction of cancer cells with uncontrollable proliferation properties. In addition, it has been reported that NOL7 functions as an mRNA-binding protein [NOL7 with respect to RNA, for instance, in RNA metabolic process and RNA processing. More importantly, KEGG pathway analysis results revealed that NOL7-related genes were mainly involved in ribosome biogenesis. Four genes that closely interacted with NOL7  which were identified in our study were implicated in ribosome biogenesis  [http://timer.cistrome.org/, accessed on 19 June 2022), based on the TCGA project [The expression data for ne 2022) . Low spe project ,27. http://gepia2.cancer-pku.cn/, accessed on 19 June 2022) [http://ualcan.path.uab.edu/analysis-prot.html, accessed on 21 June 2022), we obtained the differences in total protein expression levels of NOL7 between adjacent normal tissues and tumor tissues [NOL7 and tumor staging, we conducted a survey on UALCAN.Since several tumor tissues assessed with TIMER2.0 did not include adjacent normal tissues, we combined the data from the TCGA database with GTEx data for supplementary analyses of the expression differences between normal and tumor tissues through GEPIA2 (ne 2022) . We sele tissues . Furtherhttp://kmplot.com/analysis/, (accessed on 24 June 2022) [p-values and HRs were calculated. Moreover, we analyzed the OS and DFS of all tumors using the “survival analysis” module of GEPIA2  by setting the group cutoff to “median”. Patients were divided into a high group and a low group to generate Kaplan–Meier survival curves and survival maps and to calculate the log-rank p-values and HRs. The OS, RFS, FP, PFS, PPS, DMFS, and DSS of all tumor patients in the GEO cohorts were analyzed using the Kaplan–Meier plotter tool (ne 2022) . By settNOL7 between the various cancer tissues as compared with normal tissues. In addition, we chose the “TCGA Pan-Cancer Atlas Studies” module in the cBioPortal tool  to analyze the genetic variation in NOL7 [The UALCAN tool was used to explore the DNA methylation levels for  in NOL7 . Alterathttp://sangerbox.com/, accessed on 17 june 2022) to explore the association between NOL7 expression and immune infiltration across 33 types of tumors by choosing the “immune cell analysis (TIMER)” module. In addition, the association between NOL7 expression and many immune-checkpoint-related proteins was assessed by choosing the “immune checkpoint gene analysis” module in the same tool. Furthermore, we examined the relationship between NOL7 expression and TMB or MSI in different tumors in the TCGA cohort [p-values and partial correlation values. We used the Sangerbox tool  was used to download the top 50 NOL7-interacting molecules. We visualized the PPI network using Cytoscape [NOL7-correlated genes through GEPIA2. Pearson’ s correlation test was performed on the top five NOL7-correlated genes in the “correlation analysis” module of GEPIA2 to obtain p-values, dot plots, a heatmap, and correlation coefficients. Moreover, we generated a Venn diagram, combining the two parts using the Draw Venn Diagram tool  [p-value < 0.05 and an FDR < 0.25 were considered statistically significant.To explore the regulatory network of ytoscape . In addine 2022) . We combNOL7 in various human cancers. We propose that NOL7 may serve as a new diagnostic biomarker and therapeutic target in different tumor types, including breast cancer, colon cancer, lung cancer, and liver cancer. In addition, our work expands the understanding of the impact of NOL7 on cancer immunologic therapy, as we revealed strong correlations between NOL7 and immune cells and immune checkpoints. Future experimental and prospective studies of NOL7 in different cancer types may provide in-depth insights into regulatory mechanisms and further support the development of therapeutic strategies targeting NOL7.Overall, our first pancancer study enabled a relatively comprehensive understanding of the potential role of


HERES, TUG1, and UCA1, associated with ESCC, directly or indirectly targets various molecules of the Wnt/β-catenin pathway and facilitates the manifestation of multiple cancer phenotypes, including proliferation, metastasis, relapse, and resistance to anticancer treatment. Additionally, several lncRNAs, such as HCP5 and PTCSC1, modulate PI3K/Akt/mTOR pathways during the ESCC pathogenesis. Furthermore, a few lncRNAs, such as AFAP1-AS1 and LINC01014, block the efficiency of chemotherapeutic drugs, including cisplatin, 5-fluorouracil, paclitaxel, and gefitinib, used for ESCC treatment. Therefore, this review may help in designing a better therapeutic strategy for ESCC patients.Despite the availability of modern techniques for the treatment of esophageal squamous cell carcinoma (ESCC), tumor recurrence and metastasis are significant challenges in clinical management. Thus, ESCC possesses a poor prognosis and low five-year overall survival rate. Notably, the origin and recurrence of the cancer phenotype are under the control of complex cancer-related signaling pathways. In this review, we provide comprehensive knowledge about long non-coding RNAs (lncRNAs) related to Wnt/β-catenin and phosphatidylinositol-3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathway in ESCC and its implications in hindering the efficacy of chemotherapeutic drugs. We observed that a pool of lncRNAs, such as  Worldwide, esophageal cancer (EC) ranks eighth and sixth in terms of incidence and mortality among all cancers, respectively . DespiteIn recent years, improved knowledge of oncology research has led to the identification of non-coding RNAs (ncRNAs) that regulate tumor cell proliferation, differentiation, angiogenesis, metastasis, and invasion. ncRNAs include structural and regulatory RNAs, representing ~90% of the human genome. Structural long non-coding RNAs (lncRNAs) include ribosomal RNAs and transfer RNAs, whereas regulatory RNAs include small conditional RNAs, small nucleolar RNAs, microRNAs (miRNAs), and lncRNAs ,7. AmongLINC01014 . Fu. FuUCA1 09 cells  [11]. In pathway  , Papillary Thyroid Carcinoma Susceptibility Candidate 1 (PTCSC1), and LINC01014, and the PI3K/Akt/mTOR pathway are in tight conjunction during ESCC pathogenesis. This emphasizes the need to target this pathway with associated lncRNAs in treating ESCC patients.PI3K, Akt, and mTOR are the three significant nodes in the PI3K/Akt/mTOR pathway ,40. TyroHCP5 was upregulated in ESCC tissues compared to control tissues . At. AtLOC28d KYSE30  (Table 1 tissues . Furtherpatients  (Table 1(KYSE30) . Moreovepatients   compared with cisplatin-sensitive ESCC cells [50) value of cisplatin treatment  in ESCC cells by promoting the methylation of the promoter region of the glutathione S-transferase Pi 1 (GSTP1) gene [LINC00337 was overexpressed in ESCC patients’ tissues and cell lines, which hindered the effects of cisplatin dose  via the upregulation of TPX2 by recruiting the E2F4 transcription factor [MACC1-AS1 and FOXD2-AS1, were upregulated in ESCC cells and tumor tissues [FOXD2-AS1 increases the cisplatin resistance  by promoting the Akt/mTOR axis stimulation in ESCC cells [Furthermore, higher expression of CC cells . Howeverreatment  showed tP1) gene      by lncRN pathway . Moreoveor 48 h)  in ESCC HERES, TUG1, HCP5, and PTCSC1 and downregulated expression levels of UCA1 could be best suited for therapeutic application in clinical settings. These lncRNAs significantly target the key downstream molecules of cancer-related pathways, namely the Wnt/β-catenin and PI3K/Akt/mTOR pathway. Based on the ESCC cohort size and the detailed mechanism, UCA1, HCP5, and PTCSC1 possess great potential as a therapeutic target for ESCC in association with the Wnt/ β-catenin and PI3K/Akt/mTOR pathways, which signifies the clinical potential of lncRNAs for the treatment of ESCC patients. Thus, targeting lncRNAs and their associated pathways, i.e., Wnt/β-catenin and PI3K/Akt/mTOR, may provide novel approaches in the treatment and better management of ESCC patients.In this review, we highlight the immense potential of oncogenic and tumor suppressive lncRNAs in regulating cancer-associated signaling pathways and their implication in drug resistance in ESCC patients. As mentioned in the previous sections, the Wnt/β-catenin and PI3K/Akt/mTOR pathway comprises multiple downstream signaling proteins, such as β-catenin, GSK3-β, Akt, PI3K, and mTORC1 complex, whose activation in association with dysregulated lncRNAs can manifest several hallmarks of cancer, including uncontrolled cell growth, inhibition of apoptosis, proliferation, increased metastasis, and invasion. We found that upregulated expression levels of AFAP1-AS1 could be a selective prognostic and therapeutic marker for the ESCC patients treated with cisplatin, 5-FU, and paclitaxel. Additionally, we observed that AFAP1-AS1 lowers the efficacy of the maximum dose of cisplatin, 5-FU, and paclitaxel drugs at 24 h in 162 ESCC patients [AFAP-AS1 expression levels is of utmost importance in the management of ESCC. Furthermore, it has been shown that LINC01014 resists the efficacy of gefitinib (30 µM at 48 h), which suggests that LINC01014 requires critical attention concerning prognostic and therapeutic aspects in the clinical settings [We also presented the potential of lncRNAs as an effective regulator of chemotherapeutic drugs, such as paclitaxel, 5-FU, gefitinib, and cisplatin. In our opinion, upregulated levels of patients . Therefosettings . The divBesides, many clinical trials have shown the development of RNA therapeutics, such as miRNA mimics or antimiRs, and several are in phase II or III. Still, no lncRNA-based therapeutic agent has entered the clinical setting. In future, we believe that lncRNAs and their related signaling pathways can be targeted using secondary plant metabolites in combination with chemotherapeutic drugs for the betterment of cancer treatment, as supported by recently published studies ,65,66. EOverall, our study provides comprehensive knowledge of the lncRNA regulated Wnt/β-catenin and PI3K/Akt signaling pathways and highlights the potential of lncRNAs hindering the therapeutic efficacy in ESCC. The techniques mentioned above can be employed to target desired lncRNAs clinically. However, future studies are required to translate the findings into clinical settings.


Multi-omics expression datasets obtained from multiple public databases were used to elucidate the biological function of TK1 and its effects on clinical outcomes. The Kaplan–Meier curve, a predictive nomogram mode, and the time-dependent receiver operating characteristic (ROC) curve were established to assess the role of TK1 expression in glioma prognosis. TK1 was overexpressed in glioma compared with normal samples, and patients with elevated expression of TK1 had poor overall survival. The ROC curves indicated a high diagnostic value of TK1 expression in patients of glioma; the areas under the ROC curve (AUC) were 0.682, 0.735, and 0.758 for 1 year, 3 years, and 5 years of glioma survival, respectively. For a model based on TK1 expression and other clinical characteristics, the values of AUC were 0.864, 0.896, and 0.898 for 1 year, 3 years, and 5 years, respectively. Additionally, the calibration curve indicated that the predicted and observed areas at 1 year, 3 years, and 5 years of survival were in excellent agreement. Three types of TK1 alterations—missense mutations, splice mutations, and amplifications—were identified in 25 of 2706 glioma samples. The TK1-altered group had better overall survival than the unaltered group. Single-cell function analysis showed that TK1 was positively associated with proliferation, the cell cycle, DNA repair, DNA damage, and epithelial–mesenchymal transition in glioma. Immunoinfiltration analysis indicated that TK1 expression might play different roles in low-grade glioma and glioblastoma multiforme tumor microenvironments, but TK1 expression was positively associated with activated CD4 and Th2, regardless of tumor grade. In summary, our findings identified TK1 as a novel marker for predicting clinical outcomes and a potential target for glioma. Glioma, originating from glial or precursor cells, is the most common and deadly brain malignancy; gliomas include astrocytic tumors (including glioblastoma), ependymoma, oligodendroglioma, oligoastrocytoma, and several rare histologies . ComprehThymidine kinase 1 (TK1), a special cytosolic kinase, is involved in pyrimidine metabolism, and it fluctuates during the cell cycle ,5,6,7,8.http://gepia2.cancer-pku.cn/#index, accessed on 5 November 2021) database was used to explore differences in the mRNA expressions of TK1 genes between glioma and normal brain tissue [https://portal.gdc.cancer.gov/, accessed on 5 November 2021) tumor profiles were merged in the GEPIA2 database. We also downloaded RNA-seq and microarray expression data from the GlioVis database  [http://www.cgga.org.cn/index.jsp; accessed on 5 November 2021), and Gene Expression Omnibus . We excluded pediatric tumor samples and samples from patients who received radiotherapy, chemotherapy, or immunotherapy. We also explored TK1 expression in other cancers compared with normal samples in the Oncomine  database [The Gene Expression Profiling Interactive Analysis 2  , a powerdatabase .Using the data from the GlioVis database, we assessed the relationships ofTK1 expression with WHO tumor grade, tumor type (primary or recurrent), molecular subtype, pathological subtype, and tumor regions to determine the tumor characteristics of TK1 expression heterogeneity. Primary or recurrent tumors were determined on the basis of patient medical history. Tumor regions were defined according to magnetic resonance imaging . The conAs the GlioVis database provided detailed survival data on glioma, we assessed the relationship between glioma prognosis and TK1 expression by using the Kaplan–Meier method with the logrank test. To investigate TK1 as an independent risk factor for glioma, we further assessed the relationship of TK1 expression to age, sex, WHO tumor grade, tumor type (primary or recurrent), survival time, therapy strategy (chemotherapy and/or radiotherapy), isocitrate dehydrogenase (IDH) mutation status, and 1p19q codeletion status by using the GlioVis_CGGA data. Additionally, both the time-dependent receiver operating characteristic (ROC) curve and a predictive nomogram model were established.https://cistrome.shinyapps.io/timer/, accessed on 20 November 2021), a comprehensive website providing the molecular characteristics of tumor immune interactions [http://cis.hku.hk/TISIDB/, accessed on 29 November 2021) was used to evaluate the correlations between TK1 expression and immunoinhibitors, immunostimulators, and the abundance of tumor-infiltrating lymphocytes, chemokines, and receptors [Tumor Immune Estimation Resource  provides multidimensional cancer genomics data storage for exploring genetic alterations and their correlation with genes and clinical outcomes across samples [The cBio Cancer Genomics Portal  [DNA methylation data were downloaded from the UCSC Xena platform (n = 669) and obtained all genes coexpressed with TK1 from LinkedOmics , a publicly available portal including multi-omics data from all 32 TCGA cancer types and 10 Clinical Proteomic Tumor Analysis Consortium (CPTAC) cancer cohorts [p-value < 0.05 was deemed to indicate statistical significance.To investigate the functional mechanism of TK1 in gliomas, we selected the TCGA_GBMLGG dataset  that incorporates TargetScan, miRDB, and miRTarBase data was used to predict TK1-targeting miRNAs [We assessed the correlation between TK1 expression and miRNA expression using the microRNA_198 and mRNA-array_301 datasets from the CGGA . In thisg miRNAs . To furthttp://biocc.hrbmu.edu.cn/CancerSEA/, accessed on 10 November 2021) is a user-friendly web interface for comprehensively analyzing the gene and/or lncRNA functional states at a single-cancer-cell level [p < 0.05).CancerSEA  and human glioma cell lines (U87 and LN229) were purchased from the American Type Culture Collection. Cell lines were cultured in DMEM with 10% fetal bovine serum and 1% penicillin and streptomycin and maintained at 37 °C in an incubator set to 5% COTo confirm the protein expression of TK1 in glioma, we first reported the protein ex-pression profiles from CPTAC ,29. Of nhttp://www.R-project.org; accessed on 10 November 2022).Categorical variables were analyzed with χ2 tests and are shown as absolute counts and proportions. The Wilcoxon test and/or the Kruskal–Wallis test were used to assess non-normally distributed continuous data. A t-test was adopted to compare two continuous groups of normally distributed data, and analysis of variance was used to compare multiple groups of normal distribution data. Spearman’s rank correlation analysis was used to examine the correlation of TK1 mRNA expression with the TK1 DNA methylation value and with TK1-associated miRNA expression. The Kaplan–Meier curve was used to assess the difference in overall survival (OS) between the group with high vs. low K1 expression. The OS was calculated as the days between diagnosis and death or the end of follow-up, whichever came first. The median values for TK1 mRNA expression, TK1 DNA methylation, and TK1-associated miRNA expression were used as the cutoffs for the high and low groups. To confirm that TK1 was an independent risk factor for glioma, we used both univariate and multivariate Cox proportional hazards models in which age, sex , WHO tumor grade (II vs. III vs. IV), tumor type (primary vs. recurrent), radiotherapy (no. vs. yes), chemotherapy (no. vs. yes), IDH mutation status (wildtype vs. mutant), and 1p19q codeletion status (noncodel vs. codel) were considered. Additionally, both the time-dependent ROC and a predictive nomogram model were established. To evaluate the predictive accuracy by comparing the actual and predicted outcomes, we generated correction curves. Statistical analyses were conducted with R software , proneural (PN), mesenchymal (ME), and classic (CL) ,35. Of tIn survival analysis, the median TK1 mRNA expression value was adopted as the cutoff for the high and low groups. We identified eight datasets and then used Kaplan–Meier survival curves to assess the prognostic role of TK1 in glioma . A higheUsing the data from CGGA, we performed further univariate and multivariate Cox regression analyses, which showed that age, tumor type, WHO tumor grade, IDH mutation status, 1p19q codeletion status, chemotherapy, and TK1 expression were significantly associated with glioma prognosis, except for a broadline significant relationship between radiotherapy and glioma . We thenIn UALCAN, the protein expression of TK1 was evaluated in 10 normal tissues and 99 glioblastoma proteomic profiles from CPTAC. TK1 protein levels were also elevated in glioblastoma tissue compared with normal tissue . MoreoveUsing TIMER, we explored the associations between TK1 expression and six tumor-immune cell infiltration levels. As shown in In TISIDB, the associations between 28 tumor-infiltrating lymphocytes and TK1 expression were investigated in detail. TK1 expression was inversely associated with infiltrating levels of most kinds of lymphocytes but positively associated with activated CD4 (Act CD4) and Type 2 T (Th2) in GBM . In LGG,We also assessed the relationship between TK1 expression and three kinds of immunomodulators: immunoinhibitors , immunosWe then examined the relationship between TK1 expression and 40 kinds of chemokines (or receptors) . As showGenetic alterations of TK1 in glioma were explored with cBioPortal. Overall, three kinds of alterations, including missense mutations, splice mutations, and amplifications, were identified in 25 of 2706 glioma samples . Additiop < 0.001) between TK1 expression and TK1 DNA methylation , the cell cycle , DNA repair , DNA damage , and EMT , but negatively correlated with quiescence  and hypoxia .Using CancerSEA, we identified two GEO datasets (GSE57872 and GSE 102130) for assessing the functional characteristics of TK1 at a single-cancer-cell level in glioma. Recent studies have suggested an association between TK1 and the development of several tumors, including lung cancer, thyroid carcinoma, prostate cancer, and pancreatic cancer ,6,7,8. HIn our analysis of transcriptional data collected from TCGA, GTEx, and GEO, we discovered that TK1 expression was higher in glioma tissue samples and cell lines than in normal samples or normal cell lines. Further survival analysis indicated that patients with higher levels of TK1 expression had shorter OS. To confirm the TK1 protein expression, we assessed the expression of TK1 in normal cells and tumor cell lines. The results showed that TK1 was overexpressed in U87 and LN229 cell lines compared with SVG. Additionally, we found that missense mutations, splice mutations, and amplifications were involved in glioma samples, and our survival analysis showed that the TK1-altered group had better OS than the group with these alterations. Moreover, high TK1 expression was observed in many other tumors and was found to be associated with poor prognosis, as shown in DNA methylation is a common method of regulating gene transcription. To our knowledge, TK1 methylation has not been addressed in previous studies. For the first time, we revealed that a negative association between TK1 methylation and TK1 mRNA expression in glioma and methylation levels could determine OS in patients with glioma. Further analysis identified a similar relationship between TK1 mRNA expression and TK1 CpG sites. Notably, we did not identify the methylation levels of TK1 between tumor and normal samples; thus, further exploration is still required.p = 0.45). According to our predefined criteria, we identified that hsa-miR-1182, hsa-miR-129-5p, hsa-miR-132-3p, hsa-miR-139-3p, and hsa-miR-150-5p miRNAs may regulate TK1 expression in glioma by using the microRNA_198 and mRNA-array_301 datasets from the CGGA. Among the five miRNAs, hsa-miR-132-3p, hsa-miR-139-3p, and hsa-miR-150-5p were downregulated in glioma, and their overexpression has been found to prevent cell proliferation, colony formation, and tumor growth in gliomas [MiRNAs are small ncRNA molecules of 19–25 nucleotides that can target the 30 untranslated regions of their target mRNA to inhibit degradation and translation . Liu et  gliomas ,38,39. A gliomas . Regardi gliomas ,41. More gliomas , gastric gliomas , and non gliomas . Collectp < 0.001) in glioma, has been shown to be an oncogene functioning partially through the P53-mediated EMT signaling pathway in lung cancer cells [p < 0.001) and RRM2 , two additional top 50 TK1-associated coexpressed positive genes, inhibits the migration, proliferation, and apoptosis of glioma cells [Recent findings suggest that TK1 expression influences the proliferation of lung cancer, thyroid carcinoma, prostate cancer, melanoma, and pancreatic cancer cells ,5,6,7,8,er cells . Additioma cells ,47. TogeGrowing evidence indicates that immunological cells (both antitumorigenic and protumorigenic) in the tumor microenvironment play central roles in tumorigenesis . A previEvidence has shown that serum TK1 is a reliable biomarker for detecting malignant tumors in cancer screening ,50. In aSeveral study limitations should be acknowledged. First, we did not fully clarify the role of TK1 expression in the different pathological subtypes of gliomas because few datasets provided data on pathological subtypes, and various pathological subtypes were defined in the included datasets. As suggested in In conclusion, the current study comprehensively explored the malignant properties of TK1 in glioma through multiple levels of bioinformatics analysis. The findings provide critical insights for further investigation of TK1 as a potential target.


Fusarium graminearum is a plant filamentous pathogenic fungi and the predominant causal agent of Fusarium head blight (FHB) in cereals worldwide. The regulators of the secretory pathway contribute significantly to fungal mycotoxin synthesis, development, and virulence. However, their roles in these processes in F. graminearum remain poorly understood. Here, we identified and functionally characterized the endoplasmic reticulum (ER) cargo receptor FgErv14 in F. graminearum. Firstly, it was observed that FgErv14 is mainly localized in the ER. Then, we constructed the FgErv14 deletion mutant (ΔFgerv14) and found that the absence of the FgErv14 caused a serious reduction in vegetative growth, significant defects in asexual and sexual reproduction, and severely impaired virulence. Furthermore, we found that the ΔFgerv14 mutant exhibited a reduced expression of TRI genes and defective toxisome generation, both of which are critical for deoxynivalenol (DON) biosynthesis. Importantly, we found the green fluorescent protein (GFP)-tagged FgRud3 was dispersed in the cytoplasm, whereas GFP-FgSnc1-PEM was partially trapped in the late Golgi in ΔFgerv14 mutant. These results demonstrate that FgErv14 mediates anterograde ER-to-Golgi transport as well as late secretory Golgi-to-Plasma membrane transport and is necessary for DON biosynthesis, asexual and sexual reproduction, vegetative growth, and pathogenicity in F. graminearum. Fusarium graminearum (teleomorph: Gibberella zeae) is a devastating fungal pathogen that causes Fusarium head blight (FHB) in cereal crops, including wheat, barley, and maize [F. graminearum produces a kind of trichothecene mycotoxin named deoxynivalenol (DON), which is toxic to humans and livestock by binding to the ribosome to inhibit the protein synthesis of eukaryotic organisms [F. graminearum may facilitate the development of novel strategies and effective means for FHB control.nd maize ,2,3. FHBnd maize ,5. In adrganisms ,7. Howevrganisms ,9. ThereF. graminearum infects flowering wheat spikelets and grows through the rachis node at the bottom of each spikelet so as to colonize the entire wheat head [F. graminearum produces DON, a secondary metabolite that acts as an important virulence factor to facilitate the expansion of the pathogen in wheat heads [FHB7, which is transferred from Thinopyrum to wheat, encodes a Gluthione S-transferase and was found to catalyze DON into DON-GSH, thereby eliminating the toxicity of DON [F. graminearum, DON biosynthesis is regulated by the trichothecene (TRI) gene cluster [TRI1 and TRI4 encode cytochrome P-450 oxygenases localized on the toxisome that are involved in toxisome function [TRI5, the first gene identified in DON biosynthesis, encodes a trichodiene synthase that is essential for the first step in the DON biosynthetic pathway [TRI6 and TRI10 encode transcriptional regulators to control DON biosynthesis by regulating the TRI gene cluster expression [TRI genes, the proteins involved in vesicle transport have been reported as essential regulators of DON biosynthesis, which play critical roles in the pathogenesis of phytopathogenic fungi [Saccharomyces cerevisiae, the small GTPase Rab1 has been observed to regulate the tethering of ER-derived COPII vesicles to Golgi in vitro [F. graminearum, FgRab1 is required for virulence of pathogens [F. graminearum is the R-SNARE protein FgSec22, a subunit of the ER-to-Golgi SNARE complex, which mediates COPII-associated membrane fusion [F. graminearum [Sclerotinia sclerotiorum, SsEmp24, a protein associated with ER-derived COPII-coated vesicles, is involved in regulating pathogenicity [eat head . During at heads . Moreoveat heads . Traditiat heads . A gene,y of DON . This di cluster ,16. Amonfunction ,18,19,20 pathway ,22. TRI6pression ,24. In aic fungi ,28,29,30ic fungi . The COPin vitro ,33. In Fathogens . Anothere fusion . It has minearum . Moreovegenicity . TogetheS. cerevisiae, ER vesicle (Erv) proteins link protein cargos to subunits of COPII vesicle coat proteins and function in transporting cargo from the ER to the Golgi [S. cerevisiae that regulates the COPII cargo selection. Erv14 has been shown to link ~36 membrane proteins to COPII vesicles involved in the secretory pathway [ER-derived vesicles selectively loaded with cargo interact directly or indirectly with COPII subunits . Previouhe Golgi ,41,42. T pathway ,44,45,46 pathway , the funF. graminearum. We show that, as an ER cargo receptor, FgErv14 has important functions in DON production, vegetative growth, asexual and sexual reproduction, and pathogenicity in F. graminearum. FgErv14 is mainly located in the ER and participated in both ER-to-Golgi and late Golgi-to-plasma membrane (PM) transport. Moreover, under toxin-inducing conditions, FgErv14 is localized in the toxisome and is required for toxisome generation. Collectively, these results show that the ER cargo receptor FgErv14 is an important regulator of DON production, development, and full virulence in F. graminearum.In this work, we characterized the integral membrane protein FgErv14 in F. graminearum gene single deletion mutants were generated from the parental wild-type (WT) PH-1 strain, and all fungal strains used in this study are listed in FgERV14 disruptions, the endogenous copy of FgERV14 was replaced with a hygromycin resistance (Hyg R) cassette in WT strain PH-1. PCR and Southern blotting were used to identify ΔFgerv14 deletion mutants [FgERV14 5′ promoter region and open reading frames were amplified by PCR from the genome of F. graminearum, then, the 3′ end of the PCR product was fused with green fluorescent protein (GFP) and inserted into the pYF11 vector to generate a plasmid expressing FgErv14-GFP. The resulting pYF11-FgErv14-GFP plasmid was transferred into the ΔFgerv14 mutant protoplasts using PEG-mediated fungal transformation. The transformants were selected on a G418-resistance medium and identified by PCR and GFP signals [2O2 were used separately [The  mutants . To gene signals . Strains signals . The conparately . GenomicFor conidia germination assay, five mycelial plugs (5 mm in diameter) of all strains were taken from the edge using a 50-mm punch and induced in CMC liquid medium at 25 °C for 5 days . Then, t5 spores/mL was prepared. 10 µL of the suspension was injected into the flowering wheat spikelets. To maintain humidity of the wheat heads after injection, the ziplock bags were used after spraying with sterile distilled water. The bags were removed after 2 days, and the wheat heads showing symptoms of wheat head blight disease were collected and photographed two weeks post-inoculation. The disease index was quantified from 40 wheat heads of each strain incubation. For wheat coleoptile infection assay, 2.5 μL of three-day-old conidia suspensions (2 × 106 spores/mL) was used and the samples were photographed 8 days post-inoculation. The lesion length was quantified from 15 plants of each strain incubation.Jimai 22 was used in this assay as it was a native susceptible wheat cultivar. Infection of flowering wheat inflorescences was performed as described previously . Three-dDON biosynthesis was tested by culturing 3 fresh mycelial plugs of each strain on 5 g of wheat grains . The proTRI genes expression levels were detected using qRT-PCR with the appropriate primers and liquid trichothecene biosynthesis induction (TBI) medium induction. The FgRPS16 gene in F. graminearum served as an internal control for normalization of gene expression levels [−ΔΔCT method [Total RNA extraction was performed as previously described . The TRIn levels , and datT method .Each fungal strain expressing fluorescence-tagged proteins from plasmids and conidia stained with calcofluor white (CFW) were examined via fluorescence microscopy using confocal microscopy  or an upright fluorescence microscope  . At leasThe experimental data are displayed as the mean ± standard deviation (SD) from at least three replicates. Duncan’s test and analysis of variance (ANOVA) were performed using GraphPad Prism 8.0 to compare individual samples across different strains or treatments.F. graminearum was detected using S. cerevisiae Erv14 (YGL054C) for a homology search. The FgERV14 gene is 629 bp in length and encodes a protein of 139 amino acids. From a protein alignment analysis, it was found that FgErv14 exhibited a high degree of identity to other fungal Erv14 proteins of Fusarium oxysporum (99.28%), Magnaporthe oryzae (92.70%), Neurospora crassa (91.97%), Aspergillus nidulans (87.59%), S. cerevisiae (62.69%) and Schizosaccharomyces pombe (45.54%)  of (45.54%) . A phylo(45.54%) . Further(45.54%) . The amistrategy  and werestrategy .S. cerevisiae, the absence of Erv14, a non-essential protein, does not affect vegetative growth [F. graminearum FgErv14 in vegetative growth was investigated by comparing WT PH-1, ΔFgerv14 mutants (#3 and #4), and a ΔFgerv14/FgERV14 complemented strain cultured on V8 juice medium. The ΔFgerv14 mutant strains exhibited a phenotype of defective vegetative growth compared to that in PH-1 and the ΔFgerv14/FgERV14 strain , cell wall disruption (SDS), and oxidative stress (H2O2) revealed a significant inhibition of growth in the ΔFgerv14 strain compared to WT and ΔFgerv14/FgERV14 strains  of 12.94 and 11.89, respectively, indicating full virulence  or RabO(Rab1) blocks ER-to-Golgi transport completely, resulting in the reabsorption of early Golgi cisternae into the ER, whereas late Golgi-associated proteins were dispersed in the cytoplasm [Fgerv14 mutant, indicating that FgErv14 does not regulate the localization of FgSed5. This result is consistent with the fact that FgErv14 does not mediate all COPII anterograde transport. Given that FgErv14 has only an apparent ER localization (Fgerv14 mutant, resulting in defective late secretory transport. As secretory transport contributes significantly to the DON production, virulence, growth, and development of F. graminearum [F. graminearum.The ER cargo receptor Erv14 is highly conserved in eukaryotes, but it does not mediate the transport of all newly synthesized proteins. The golgin protein Rud3, the general amino acid permease Gap1, and the Nast Erv14 ,56,67. Iminearum A, demonso the PM B. Accordytoplasm ,61,68. Olization A, we speminearum ,69, FgErF. graminearum. FgErv14 was observed in the toxisome, where it plays an important role in toxisome generation. We also found that FgErv14 participates in ER-to-Golgi transport and thus, in the regulation of F. graminearum virulence.In summary, our study demonstrated the involvement of the ER cargo receptor FgErv14 in vegetative growth, DON production, asexual and sexual reproduction, and pathogenicity in