The most frequently used epigenetic drugs were histone deacetylase inhibitors followed by antisense oligonucleotides, DNA methyltransferase inhibitors and histone demethylase inhibitors, the last of which are only being tested in prostate cancer. In more than 50% of the clinical trials considered, epigenetic drugs were used as part of combination therapy, which achieved the best results. The epigenetic regulation of some cancers is still matter of research but will undoubtedly open a window to new therapeutic approaches in the era of personalized medicine. The future of therapy for urological malignancies is likely to include multidrug regimens in which epigenetic modifying drugs will play an important role.Epigenetic dysregulation is one of many factors that contribute to cancer development and progression. Numerous epigenetic alterations have been identified in urologic cancers including histone modifications, DNA methylation changes, and microRNA expression. Since these changes are reversible, efforts are being made to develop epigenetic drugs that restore the normal epigenetic patterns of cells, and many clinical trials are already underway to test their clinical potential. In this review we analyze multiple clinical trials ( Urologic cancers account for approximately 10% of all cancer deaths in the USA and include bladder, kidney, prostate and testicular cancers [The establishment and progression of malignancy involves broad changes in gene expression that are determined by both genetic and epigenetic events. Genetic events include chromosome rearrangements and duplications as well as translocations, deletions, and single base pair mutations. Epigenetic modifications are somatically heritable changes that modify gene expression without altering the DNA sequence. Among these are histone modifications, DNA methylation, and miRNA expression .Post-translational modification of the histone protein N terminal tails can alter the structure of the nucleosome and change the compaction state of chromatin. Common modifications include methylation, acetylation, phosphorylation, ubiquitylation and sumoylation . Among tHistone acetylation neutralizes the positive charge of lysine residues, weakening their electrostatic interactions with DNA . This leHistone methylation occurs at lysine and arginine residues and, depending on the target, can lead to activation or repression of gene expression . MethylaIn cancer, histone modifications have been associated with both activation and repression of gene expression. Modifications such as histone 3 methylation at lysine 4 (H3K4me), histone 3 di-methylation at lysine 4 (H3K4me2), histone 3 tri-methylation at lysine 4 (H3K4me3), histone 3 acetylation at lysine 9 (H3K9ac), histone 3 methylation at lysine 9 (H3K9me) and histone 3 acetylation at lysine 27 (H3K27ac) are associated with active chromatin whereas histone 3 tri-methylation at lysine 36 (H3K36me3), histone 3 tri-methylation at lysine 9 (H3K9me3) and histone 3 methylation at lysine 27 (H3K27me) are associated with repressive chromatin [DNA methylation results from addition of a methyl group to the 5-carbon of a cytosine residue by the enzyme DNA methyltransferase (DNMT) . DNMT fode novo methylation [In eukaryotes, DNA methylation is mediated by three DNMTs: DNMT1 is responsible for the maintenance of methylation patterns after DNA replication whereas DNMT3A and DNMT3B carry out hylation . Any altmiRNAs are small endogenous non-coding RNAs (ncRNAs), 21-25 nucleotides in length, that regulate gene expression by targeting specific messenger RNAs (mRNAs) for translational repression or degradation. Expression patterns of miRNAs differ between normal and tumor tissues , 17. DepTwo strategies for epigenetic therapy are currently in use: small molecules that inhibit epigenetic-modifying enzymes and manipulation of miRNA expression.Amongst the small molecule inhibitors are HDAC inhibitors and DNMT inhibitors. HDAC inhibitors (HDACi) are classified into 4 groups according to their chemical structures: hydroxamates , cyclic peptides (Romidepsin), benzamides (Entinostat and Mocetinostat) and aliphatic fatty acids  .The majority of HDACi inhibit zinc-dependent HDACs by interacting with the zinc ion. In cancer cells, the inhibition of histone deacetylation restores expression of tumor suppressor genes that were previously silenced by epigenetic mechanisms , 19.DNMT inhibitors are divided into nucleoside analogues and non-nucleoside analogs . NucleosAnother focus of epigenetic therapy is the manipulation of miRNA expression and activity. Several strategies have been employed to silence miRNAs that are overexpressed in cancer. These include anti-miRNA oligonucleotides (AMOs), peptide nucleic acids (PNAS), miRNA-masking antisense oligonucleotides (miR-mask) and miRNA sponges . RestoraDysregulation of epigenetic marks leads to changes in gene expression that, in cancer cells, can result in activation of oncogenes or inactivation of tumor suppressor genes, both of which can contribute to cancer. Unlike genetic mutations, however, epigenetic changes are reversible. Therefore, the development of drugs capable of restoring the normal epigenetic patterns of cells has great therapeutic potential. In this review we discuss the efficacy of this novel therapeutic approach through the analysis of clinical trials of epigenetic therapies conducted in prostate, kidney and bladder cancers.https://clinicaltrials.gov/) and European  databases. Relevant articles on the subject were also retrieved from PubMed database using keywords encapsulating all types of epigenetic therapies and urologic cancers . To guarantee that most of the data on the subject was included, the reference sections of the captured articles were also filtered for relevant articles.We performed a comprehensive literature review and searched for clinical trials from the United States . Currently there are several chemotherapeutic agents approved for the treatment of advanced CRPC: Sipuleucel T, Docetaxel, Cabazitaxel, Abiraterone, Alpharadin and Enzalutamide –35. Althet al showed that low doses of the HDACi Panobinostat can restore the susceptibility of prostate cancer cells to hormonal therapy with the nonsteroidal antiandrogen Bicalutamide [in vivo in the context of hormonal therapy [Pre-clinical studies in prostate cancer cell lines demonstrate that treatment with HDACi can restore susceptibility to chemotherapeutic agents such as taxanes, antiandrogens, and mTOR inhibitors –40. Comblutamide . The HDA therapy .in vitro study of the HDACi Panobinostat in combination with the mTOR inhibitor Rapamycin in prostate cancer cell lines resulted in a decrease in HIF1- α expression leading to inhibition of angiogenesis [An ogenesis . Combineogenesis .et al revealed that prostate stem-like cells are more resistant to radiotherapy [Cancer stem cells are thought to be responsible for treatment resistance and tumor recurrence  and presotherapy . Howeverotherapy . Additiootherapy .Cancer is a heterogeneous disease and the identification of biomarkers that predict whether a specific therapy (including epigenetic therapies) will be beneficial, is essential to improving cancer treatment. Recently, it has been reported that prostate cells positive for the presence of androgen receptor and cellular prostatic acid phosphatase show greater response to treatment with HDACi than cells without this pattern of expression .GSTP1 methylation levels and a consequent increase in gene expression [In vivo, treatment of immunodeficient mice carrying xenograft tumors with Procainamide resulted in a significant reduction in tumor size, suggesting clinical efficacy [At the level of DNA methylation, reversion of methylation can restore expression of genes silenced by this epigenetic mechanism. Treatment of human prostate cancer cells with Procainamide, a non-nucleoside DNMT inhibitor, results in a decrease in pression . In vivoefficacy .In vivo studies reveal that long-term treatment of prostate cancer cells with the DNMTi Azacitidine led to a significant reduction in cell proliferation due to increased androgen receptor expression. Moreover, androgen receptor induction restored sensitivity to the antiandrogen agent Bicalutamide [Resistance to hormonal therapy in prostate cancer is mediated by several mechanisms. Alterations at the DNA level include androgen receptor gene amplifications and point mutations . Howeverlutamide .APC and RARβ gene expression, both of which are known to be hypermethylated and inactive in prostate cancer [in vitro, and in vivo using prostate cancer xenograft models [The DNA demethylating agent, Disulfiram, has also been tested in prostate cancer cells. Treatment with Disulfiram resulted in the reestablishment of e cancer . Cell grt models .CDK1, CDK2 and BCL2 [In vivo, Takeshita et al used the atelocollagen method to deliver miR-16 to bone metastases via the mouse tail vein. They subsequently observed a suppression in metastasis growth, indicating not only efficacy of the treatment but also of the delivery method [Finally, microRNAs modulators have been tested in preclinical studies as potential therapeutic options for prostate cancer. miR-16 regulates the expression of genes involved in cell-cycle control and apoptosis such as and BCL2 , 50. Traand BCL2 . In vivoy method .Bcl2 and CLU genes in prostate cancer is linked to chemoresistance and cancer progression [HDAC1 using an siRNA [Like HDACi and DNMTi, antisense oligonucleotides can restore the sensitivity of cancer cells to chemotherapeutic agents. Upregulation of the gression , 52. Knogression , 52. Furan siRNA . The inhan siRNA .These results demonstrate the potential for epigenetic therapies to advance prostate cancer treatment.Clinical trials using HDAC inhibitors for the treatment of prostate cancer showed a PSA response in five studies, three of which resulted in a decrease in PSA levels of ≥50%  inhibitor used in the treatment of psychiatric disease. MAOA is an enzyme responsible for the deamination of neurotransmitters, such as dopamine, serotonin and norepinephrine, that are important in a variety of neurological and psychiatric illnesses . MAOA haIn the area of miRNA modulation, four clinical trials of antisense oligonucleotides have reported a positive PSA response, with three trials describing a PSA response >=50%  by either genetic and epigenetic factors [VHL inactivation in both sporadic and familial forms can occur due to point mutations or deletions at the genetic level, or due to DNA hypermethylation at the gene promoter [Clear cell RCC is the most common form of renal cell carcinoma, and is associated with inactivation of the tumor suppressor gene  factors , 77. VHLpromoter , 77.RASSF1A and HIC in patients with RCC is associated with a poor prognosis [DNA methylation also affects the regulation of several genes in RCC and has the potential to be used as biomarker and as a therapeutic target . For exarognosis , 80.Several miRNAs showed altered expression in RCC, resulting in changes to important cellular functions such as apoptosis, angiogenesis and the epithelial mesenchymal transition , 81. ExaRadical nephrectomy is the standard of care for localized renal cell carcinoma. However, high rates of recurrence after surgery demand the development of new adjuvant therapies. Both radiotherapy and hormone therapy have proven ineffective in advanced stages of disease and chemotherapy has a response rate inferior to 10% , 83.Preclinical studies using epigenetic drugs for kidney cancer treatment show some promise. In renal cancer cell lines, the HDACi Panobinostat induced cell cycle arrest and apoptosis and also resulted in a reduction in tumor size in xenograft mice models . Pre-cliOne ongoing issue in RCC treatment is resistance to immunomodulatory therapy with interferons. This can occur via promoter hypermethylation and silencing of interferon response genes . TreatmeVHL cell status with lower levels of miR-30c being observed in VHL-deficient RCC cell lines [Downregulation of miR-30c in RCC is associated with promotion of the epithelial mesenchymal transition . Reducedll lines . Transfell lines .Trials of HDACi in RCC have shown mixed responses. HDACi Vorinostat used as a monotherapy showed an objective response in 36% of patients, however 63% of the patients presented disease progression at 6 months Table : s. WhenThe only DNMT inhibitor tested in RCC is Decitabine. When used in combination with the cytokine IL-2 in a phase II study of advanced RCC, three out of five patients showed stable disease . Anotheret al, in which the antisense oligonucleotide MG98 was administered to 15 patients, no objective responses were observed with nine patients presenting progression of the disease. MG98 targets DNMT1 but no decrease in enzyme activity was observed [Treatment with antisense oligonucleotides has also resulted in stabilization of disease in some RCC patients. The antisense oligonucleotide GTI-2040, targeting the R2 subunit of ribonucleotide reductase, was tested in patients with metastatic disease in a phase II trial and generated a partial response for approximately eight months . One patobserved . Also, gobserved .miRNA mimics have also been trialed in RCC. MRX34, an miRNA mimic of the tumor suppressor miRNA34, was tested in a phase I clinical trial for advanced or metastatic cancers, including RCC, but the trial was terminated early due to serious immunologic adverse events  avoid some of the issues described above as as they are designed to be more target specific. Thus far, two separate studies of ASOs as monotherapy have shown no objective responses , 94, howCurrently, cancer treatment is determined largely according to cancer stage, even though patients with similar stage cancers may respond differently to the same type of therapy. The promise of “personalized medicine” is the idea that tailoring treatment to an individual patient will optimize efficacy while minimizing toxicity. Personalization should be an ongoing goal for all cancer therapy development, including epigenetic therapies. Biomarker development should thus be a central goal in the development of epigenetic therapies, both so that the correct patient receives the correct therapy, and to ensure that therapies that have value in a subset of patients are not passed over because of lack of efficacy in other patients. Epigenetic therapies are still in their infancy as a therapeutic class and pre-clinical promise has not yet translated into clinical efficacy. However, the development of target-specific agents, and the careful combination of epigenetic therapies with traditional modalities should enable them to achieve clinical success in the near future.


The centromere drive model explaining rapid evolution of eukaryotic centromeres predicts higher frequency of positive selection acting on centromeric histone H3 (CenH3) in clades with asymmetric meiosis compared to the clades with only symmetric meiosis. However, despite the impression one might get from the literature, this key prediction of the centromere drive model has not only never been confirmed, but it has never been tested, because all the previous studies dealt only with the presence or absence instead of the frequency of positive selection. To provide evidence for or against different frequencies of positively selected CenH3 in asymmetrics and symmetrics, we have inferred the selective pressures acting on CenH3 in seventeen eukaryotic clades, including plants, animals, fungi, ciliates and apicomplexa, using codon-substitution models, and compared the inferred frequencies between asymmetrics and symmetrics in a quantitative manner. We have found that CenH3 has been evolving adaptively much more frequently in clades with asymmetric meiosis compared with clades displaying only symmetric meiosis which confirms the prediction of centromere drive model. Our findings indicate that the evolution of asymmetric meiosis required CenH3 to evolve adaptively more often to counterbalance the negative consequences of centromere drive. Centromeric histone H3 (CenH3) is the cornerstone of the kinetochore, which ensures the proper segregation of chromosomes during cell division by mediating chromosomal attachment to spindle microtubules. This conserved function implies that the kinetochore should be evolving under strong purifying selection  over evolutionary time, but multiple reports have suggested that CenH3 has been evolving adaptively  in various lineages of plants and animals1234567991011129999Plasmodium and two species of Ostreococcus have been analyzed151615The prediction that follows is that adaptively evolving CenH3 should be more frequent in clades with asymmetric meiosis than in clades with only symmetric meiosis, where there is no opportunity for conflict because all four meiotic products survive9Taken together, the centromere drive model predicts higher frequency of positive selection episodes in the evolution of CenH3 in eukaryotic clades with asymmetric meiosis (from now on referred to as asymmetrics) than in clades with only symmetric meiosis (from now on referred to as symmetrics). Since not just centromere drive, but other factors may affect the incidence of positive selection events and thus mask the effects of centromere drive, it is necessary to analyze a large enough and representative dataset to detect statistically significant difference in positive selection frequency between the asymmetrics and symmetrics. To achieve such representativeness, we analyzed 191 CenH3 sequences  sequences from a total of seventeen eukaryotic clades of plants, animals, fungi, ciliates and apicomplexa differing in meiotic symmetry  the proportion of positively selected branches in the tree, (ii) the proportion of positively selected codon in the alignments and (iii) the overall ω ratio. To determine the frequency of positively selected branches, we ran the data through the branch-site random effects likelihood (BS-REL) model of codon substitution24pgls) as implemented in caper R-package , all of them from the asymmetrics, while no positively selected branches were detected in the symmetrics , which is in accordance with the prediction of the centromere drive model. However, such a result does not rule out positive selection from symmetrics because the corrections for multiple testing may be too conservative and may lead to false-negative results. Therefore, we have also used uncorrected p-values and compared the proportions of positively selected branches between asymmetrics and symmetrics  in asymmetrics and symmetrics, we analyzed CenH3 from each of the seventeen clades using MEME (see Methods for details). The symmetrics had a significantly lower proportion of positively selected codons than asymmetrics (pTrichoderma) of nine clades (22%) showed positively selected codon in the HFD, while in asymmetrics, five of eight (63%) displayed positively selected codons in this domain  compared with clades without such opportunities (symmetrics). In accordance with the predictions of the centromere drive model and previous reports based on smaller datasets1516Tetrahymena from asymmetrics , centromeric DNA has been rapidly evolving due to very high mutation rate3416Saccharomyces due their symmetric meiosis, these findings suggest that meiotic asymmetry may indeed be the main factor responsible for the adaptive evolution of CenH3.A recent study suggested that the evolution of centromeric DNA repeats and CenH3 might be driven by selection of centromere-linked genes rather than by centromere drive in inbred lines of maizeLuzulaIn the light of the centromere drive model and the results of our and previous studies, it is likely that before asymmetric meiosis appeared on the evolutionary scene, CenH3 had been evolving under strong purifying selection with occasional episodes of positive selection. After meiotic asymmetry appeared, positive selection occurred at a higher frequency because CenH3 was forced to counteract the negative consequences of centromere drive. Interestingly, some asymmetrics have evolved holokinetic chromosomes10How to cite this article: Zedek, F. and Bureš, P. CenH3 evolution reflects meiotic symmetry as predicted by the centromere drive model. Sci. Rep.6, 33308; doi: 10.1038/srep33308 (2016).


Saccharomyces cerevisiae to human cell lines and primary cells. Emodin binds to redox-active enzymes and its effectiveness depends on the oxidative and respiratory status of cells. We show that cells with efficient respiratory metabolism are less susceptible to emodin, whereas cells under glycolytic metabolism are more vulnerable to the compound. Our findings indicate that emodin acts in a similar way as known uncouplers of the mitochondrial electron transport chain and causes oxidative stress that particularly disturbs cancer cells.The anthraquinone emodin has been shown to have antineoplastic properties and a wealth of unconnected effects have been linked to its use, most of which are likely secondary outcomes of the drug treatment. The primary activity of emodin on cells has remained unknown. In the present study we demonstrate dramatic and extensive effects of emodin on the redox state of cells and on mitochondrial homeostasis, irrespectively of the cell type and organism, ranging from the yeast  Emodin  is a naturally occurring anthraquinone present in a wide variety of plants, particularly in roots and barks , anti-fuOne of the main characteristics of cancer cells compared to healthy ones is an extensive prooxidative state that can lead to intrinsic oxidative stress . ElevateThis work originates from the observation that emodin affects cancer cells in culture while healthy cells are apparently resistant to its effects. To unravel the molecular mechanism of the compound, we performed targeted and global proteomics analyses of healthy and cancer cells treated with emodin . We charEmodin has a detrimental effect on cancer cells in culture, while healthy cells apparently remain unaffected by the compound. In this study, we evaluated the response of primary normal human fibroblasts from skin (NHF), normal human keratinocytes (NHK), and of MCF7, A549, CaCo-2 and HeLa cells, which are cancer cell lines isolated from breast, lung, colon and cervix carcinomas, respectively, to emodin. Figure To study the molecular basis of decreased cell proliferation by emodin treatment we analyzed signaling pathways known to play a role in cell proliferation and growth. Analysis of mechanistic target of rapamycin (MTOR) kinase activity by measuring the levels of phosphorylation of its direct target ribosomal protein S6 kinase revealed a perturbation by emodin, in correlation with decreased cell proliferation and protein synthesis Figure . It is e+ and NADPH -based quantitative mass spectrometry (MS) of CaCo-2 cells was employed to identify proteins binding to emodin-coated sepharose beads compared to ethanolamine-blocked control beads Figure  14]. In. In14]. H Figure .2O2 levels and detected an increase in all cells treated with emodin  were mostly unaffected , and of the citric acid cycle  appeared unaffected. Levels of marker proteins of the mitochondrial outer and inner membrane were unaltered, as well. Classical effectors of mitochondrion-associated apoptosis  are incapable to perform respiration, due to the lack of mitochondrial DNA-encoded essential subunits of the respiratory chain, and therefore solely depend on fermentation. ρ0 cells use ATP to maintain the basal mitochondrial membrane potential, which is a prerequisite for essential mitochondrial functions, such as Fe/S cluster biosynthesis [0 cells are still able to undergo mitochondrion-dependent cell death [+ cells) can switch their metabolism between fermentation and respiration dependent on the availability of the main carbon sources. In medium with glucose (Glc) as carbon source yeast cells undergo fermentation, mainly because of catabolite repression of respiratory enzymes by the presence of glucose [0 cells grown on glucose containing media (Glc) (no respiratory capacity), ρ+ cells grown on glucose containing media (Glc) , and ρ+ cells grown on galactose/lactate -containing media (high respiratory capacities). Yeast cultures treated for one to four days with emodin were evaluated for levels of oxidative stress, which can be detected by the intracellular conversion of the ROS-sensitive stain dihydroethidium (DHE) to fluorescent ethidium [0 strain under Glc conditions). A highly active and efficient mitochondrial respiratory chain protects from emodin , whereas cells with an intact respiratory chain but that primarily undergo fermentation  are highly sensitive to emodin. Thus, the respiratory capacity of yeast cells determines the cytotoxicity of emodin.With the aim of further studying the sensitivity of cells with different respiratory capacities to emodin, we employed the yeast th media . Yeast cynthesis  and protynthesis . Consequll death , and to ll death , 28. In  glucose . In contethidium . This ap+ strain, Glc) with different concentrations of the antioxidant NAC, and measured the levels of ROS and yeast cell survival. We observed that increasing the concentration of NAC reduced the levels of oxidative stress in cells treated with emodin and increased the levels of oxidative stress in control cells . Emodin strongly increased oxygen consumption of isolated yeast mitochondria in the presence of respiratory substrates Figure . AdditioIt is known that the physiological status of mitochondria in cancer cells differs from that in healthy controls. We evaluated the influence of emodin on the mitochondrial activity of healthy NHF, and as examples for cancer cells on A549 and HeLa cells to characterize the extent to which these cells were affected by the compound Figure . InteresEmodin proved to affect mammalian cells in a differential manner, dramatically interfering with cancer cell viability, without noticeable consequences for healthy cells. With the aim of understanding the molecular mechanism by which emodin causes these differential effects, we based our studies on targeted and unbiased MS-based chemoproteomics. We identified emodin binding partners and in a global fashion analyzed proteomic changes of cancer and healthy cells upon emodin treatment. Emodin interferes with the cellular redox system by binding to NADPH-dependent oxidoreductases. In addition, in all tested cells emodin significantly affects levels of mitochondrial proteins, particularly of subunits of the respiratory chain complexes. The clearest effect of emodin is the marked decrease in complex I proteins, which is observed more effectively in cancer cells compared to healthy ones. All other electron transport chain complexes show lower protein levels after emodin treatment, however to a lesser extent than complex I.0 yeast cells to be more resistant to emodin treatment, since they do not rely on mitochondria to generate ATP. However, mitochondria perform key cellular functions other than respiration. Also ρ0 cells, which cannot respire, do have a (reduced) mitochondrial membrane potential, largely generated by electrogenic transport processes, like the uptake of ATP from the cytosol in exchange with ADP generated in the mitochondrial matrix by the uncoupling hydrolytic activity of the F1-ATPase. Mitochondria are functionally important in these cells for other metabolic processes including Fe/S cluster assembly [0 cells through its uncoupling activity is immediately adverse to their survival.The yeast results are particularly interesting since they demonstrate that the respiratory capacity of cells is of critical importance for the cell's susceptibility to emodin. The compound particularly damages cells kept in fermentative growth conditions. Increasing the respiratory capacity of yeast cells protects them from the lethal effect of emodin. Along these lines, it seems intuitive to expect ρassembly . It is pAlthough the Warburg effect is as yet a matter of discussion, it is accepted that cancer cells rely more on glycolysis than on respiration to obtain energy. As seen from the experiments in yeast, emodin specifically affects cells with a strong fermentative metabolism, but spares cells with high respiratory rates. Thus, cells with high respiratory capacity, like healthy cells, are protected from emodin, whereas cells with less respiratory capacity, as cancer cells, are sensitive to emodin. In well agreement, pre-incubation of mammalian cells with DOX, known to affect 28S mitochondrial ribosomal subunits, renders healthy cells more sensitive to emodin treatment, whereas cancer cells are unaffected by the additional DOX treatment. The marked fragmentation of mitochondria and the strong decrease of mitochondrial proteins in treated cells suggest additional consequences on mitochondria other than the decrease in complex I proteins. As observed by MS, the depletion of the mitochondrial fusion protein OPA1 and of the protease YME1L1 correlates to the observed mitochondrial fragmentation upon emodin treatment. Of note, it has been shown that dissipation of Δψ induces enhanced proteolytic cleavage of OPA1 by the protease OMA1 leading to the fragmentation of the mitochondrial network –35.2O2, which is also an important signaling molecule, to highly toxic chemicals such as superoxide anions or hydroxyl radicals. We demonstrated increased H2O2 levels, but cannot rule out that the levels of other ROS are also altered. Of note, respiratory chain-derived ROS are primarily superoxide anions that are subsequently converted to H2O2 [The direct protein targets of emodin proved to be redox-active enzymes. Nevertheless, our data suggest that another critical molecular mechanism of emodin action is the partial uncoupling of mitochondria. As the depletion of the membrane potential occurs immediately after the addition of emodin to isolated mitochondria, judged by analysis of membrane-potential-dependent preprotein import and oxygen consumption rates of mitochondria Figure , this ef to H2O2 . The uncAltogether, the majority of the published effects of emodin appear to be of secondary nature. This comprehensive study identifies emodin as binding to NADPH-dependent redox-active enzymes and as a mitochondrial uncoupler, decreasing mitochondrial membrane potential. These alterations lead to an increased oxygen consumption and elevated oxidative stress levels causing a decrease in the levels of complex I , possiblAdditional Materials and Methods can be found online in supplementary information.Cells were cultured in DMEM (high glucose) supplemented with 10% fetal calf serum, penicillin/streptomycin , and 2 mM l-glutamine. For the labeling, cells were cultured in SILAC-DMEM supplemented with 10% dialyzed fetal calf serum, penicillin/streptomycin , 2 mM l-glutamine, 42 mg/L l-arginine 13C6-15N4, 73 mg/L l-lysine 13C6-15N2 (Arg10-Lys8), and 82 mg/L proline.5 cells were seeded per 10 cm plate and harvested the next day using a cell scraper. Harvested cells were lysed with 80 μL of SDS lysis buffer, incubated for 20 min with benzonase, and centrifuged at 16.000 rcf for 15 min. Supernatant was used for determination of protein concentration employing the Thermo Scientific Pierce BCA Protein Assay Kit as recommended by the supplier.8 × 10CellROX® Green Reagent from ThermoFisher Scientific was used as recommended by the manufacturer on live cells.


Legionella pneumophila, is restricted by the retrograde trafficking pathway. L. pneumophila employs the Icm/Dot type IV secretion system (T4SS) to form the replication-permissive Legionella-containing vacuole (LCV), which is decorated with multiple components of the retrograde trafficking machinery as well as retrograde cargo receptors. The L. pneumophila effector protein RidL is secreted by the T4SS and interferes with retrograde trafficking. Here, we review recent evidence that the LCV interacts with the retrograde trafficking pathway, discuss the possible sites of action and function of RidL in the retrograde route, and put forth the hypothesis that the LCV is an acceptor compartment of retrograde transport vesicles.Retrograde trafficking from the endosomal system through the Golgi apparatus back to the endoplasmic reticulum is an essential pathway in eukaryotic cells, serving to maintain organelle identity and to recycle empty cargo receptors delivered by the secretory pathway. Intracellular replication of several bacterial pathogens, including  Eukaryotic cells employ complex endomembrane systems to process internalized material and transport newly synthesized proteins from the endoplasmic reticulum (ER) via the Golgi apparatus to target organelles and membranes  and for receptor recycling between endosomes and the plasma membrane (PM)  hydrolyze PtdInsP2 and PtdInsP3 to yield PtdIns(4)P and PtdInsP2  is required for retrograde trafficking from EEs to the Cargo shuttled via the retrograde endosome-to-TGN pathway comprises many different membrane and soluble proteins as well as lipids. Membrane protein cargos include Vps10 domain family cargo receptors such as cation-dependent or cation-independent mannose 6-phosphate receptor (CD- or CI-MPR) and sortilin/Vps10  and different combinations of sorting nexin (SNX) proteins, which harbor a PI-binding PX (phox homology) domain  domain  proteins are necessary for selective delivery to target membranes  . This T4SS injects more than 300 different “effector” proteins into the host cell, dictating every step from uptake to interference with various vesicle trafficking pathways and autophagy, interaction with the ER, and finally escape from the host cell  and for downstream events leading to fusion of the phagosome with bactericidal lysosomes P again accumulates on LCVs in an Icm/Dot-dependent manner, preceding attachment of the ER  was identified as a binding partner of the retromer complex, and the effector protein promotes intracellular bacterial replication P, respectively. Ectopically produced RidL inhibits retrograde trafficking of fluorescently labeled STxB in HeLa cells, as evidenced by reduced Golgi localization of the toxin. Furthermore, in macrophages infected with wild-type L. pneumophila, retrograde trafficking of fluorescently labeled CTxB is impaired in a RidL-dependent manner. In presence of RidL, CTxB colocalizes with an EE/RE marker (transferrin receptor), but not with markers for lysosomes (dextran) or the Golgi apparatus (GM130). In absence of RidL, however, the toxin colocalizes with both EE/RE and Golgi markers, but not with the lysosome marker, consistent with a RidL-dependent inhibition of retrograde trafficking at endosome exit sites  adopts a new “foot-like” fold comprising a protruding hydrophobic β-hairpin at its “heel,” which interacts with Vps29  and CI-MPR decorate a larger portion of LCVs. Moreover, the late endosomal/lysosomal marker LAMP1 increasingly localizes to LCVs in the absence of RidL. A larger portion of LCVs also stain positive for SNX1 and SNX2 in the absence of RidL, possibly because RidL competes with the SNXs for PtdIns(3)P binding. Additionally, the level of the retrograde trafficking regulator and Rab7 GAP TBC1D5 is increased on LCVs in absence of RidL  and represent multi-domain proteins with distinct  activities  activities. Indeed, several in cis (on the LCV) but might also function in trans (in a distance from the pathogen vacuole). Most available data is in agreement with the pathogen vacuole being an acceptor compartment in the retrograde trafficking pathway, such that RidL blocks the productive interaction of the LCV with incoming retrograde vesicles in cis, rather than inhibits the formation of retrograde transport vesicles at endosome exit sites in trans  as well as the retromer subunits and the bacterial effector localize to LCVs. Yet, the retrograde toxin cargos STxB or CTxB cannot be detected on the LCV membrane, although their trafficking is affected by RidL. Due to its presence on the pathogen vacuole, RidL seems to act s Figure . FurtherAll authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Conciliospongia anjiensis gen. et sp. nov., from the Late Ordovician (~444 Ma) Anji Biota of South China. This species has a reticulate, tufted skeleton of minute monaxon spicules, characteristic of the fossil demosponge family Hazeliidae and modern heteroscleromorphs, with hexactine spicules and a globose body form inherited from reticulosan ancestors. This transitional morphology had previously been hypothesized in palaeontological studies. This morphological intermediate between two extant classes further confirms siliceous sponge monophyly and demosponge–hexactinellid spicule homology, and supports the primitive, stem-silicean interpretation of simpler-structured fossil reticulosans.The two major extant groups of siliceous sponges, Demospongiae and Hexactinellida, are generally regarded as sister groups forming the clade Silicea, although the nature of their last common ancestor is uncertain. The fossil record contains a diverse range of basal demosponges that appear to have evolved from hexactine-bearing reticulosan ancestors, although a compelling morphological intermediate has not previously been discovered. Here we describe a new species of fossil sponge,  This record, however, consists primarily of those groups with rigid, lithistid skeletons, or hypercalcified groups6. The skeleton of other demosponges is either primarily organic1, sometimes with some silica and/or aragonite biomineralization associated with chitinous fibres7, or consists of small siliceous spicules1 that are difficult to preserve articulated. These spicules can be relatively large in various groups of tetractine-bearing demosponges, but this appears to be a derived condition8; in the majority of Heteroscleromorpha2, such as the haplosclerids9, the primary spicule type is minute (sub-millimetre) monaxons, with even smaller microscleres.Demosponges are the most diverse and abundant group of extant sponges4. Most of the taxa from Cambrian rocks that have been traditionally described as protomonaxonid demosponges10 are now recognized as belonging to a much earlier-branching group of sponges12. Keratosan sponges have been described only occasionally, with the best-known examples being the family Vauxiidae, primarily from the Cambrian Burgess Shale-type faunas15. A few later Palaeozoic examples of keratosans are also known18, together with some agglutinating taxa that probably also fall into the Keratosa or Verongida19. Spiculate demosponges have been much harder to identify and interpret accurately, and most early examples are included in the Hazeliidae14. This family is characterized by minute monaxon spicules comprising a normally reticulating array of small overlapping tufts, and have been interpreted as basal crown-group demosponges12. In addition, an early Cambrian disarticulated heteroscleromorph has been described, showing the origin of their typical microscleres within the clade20; microscleres appear to have been absent from hazeliids.The fossil record of non-lithistid, non-hypercalcified demosponges is relatively sparse23, including some acceptance of the likely homology of siliceous spicules between the two classes24. Other topologies, however, are still being occasionally recovered25. The explicit question of homology of spicules between demosponges and hexactinellids is normally avoided, although significant differences in secretion mechanism and structure have been noted27. Molecular analysis8 left open the question of whether demosponge skeletons were primitively spicular or organic, but palaeontological revision of the hazeliids and vauxiids12 implies spiculate ancestors, with spicules also present in basal vauxiids; the presence of silica in vauxiids has since been confirmed15. Furthermore, a study of spicule structure in the Ordovician hexactine-bearing reticulosan Cyathophycus28 demonstrated hexagonal axial canal symmetry diagnostic of demosponges. This supports a speculative earlier suggestion29 that demosponges may have arisen from a sponge similar to Cyathophycus through loss of the external hexactine layer and exaggeration of the inner layer of fine monaxons. The new Ordovician sponge described herein . Conciliospongia therefore illustrates the stem group of demosponges, at a stage prior to the complete loss of the primary symplesiomorphies  and also before the origination of shared demosponge characters .The new species precisely fills this gap in the theoretical sequence. The globose body form is identical to that of 30 is relatively late (Late Ordovician) in comparison with previously-described early Cambrian crown-group demosponges20, indicating a significant period of evolution since the separation of this sponge from the lineage leading to the base of the demosponge crown group , a roadside section near the Fushi Reservoir, Anji County. The material can be dated as late Metabolograptus persculptus Biozone  on the basis of graptolite biostratigraphy30. Material was photographed using a Zeiss Discovery V.20 stereomicroscope and Axiocam 506 camera system (photomicrographs), or with Nikon D80 with extension tubes and 105 mm macro lens (wider-angle photographs). Specimens are deposited in the Nanjing Institute of Geology and Palaeontology (NIGP).The studied specimens were recovered as part of a diverse sponge fauna from the Upper Ordovician Wenchang Formation in Zhejiang Province, China


Notwithstanding, the imbalance of Ca2+ concentrations, altered signaling pathways, induction of apoptotic factors, and heart remodeling leading to abnormal anatomy have already been reported. Of note, a misbalance of signaling biomolecules, such as kinases and tumor suppressors , seems to participate in apoptotic and fibrotic events. In HCM, structural and cellular information about defective sarcomeric proteins and their altered interactome is emerging but still represents a bottleneck for developing new concepts in basic research and for future therapeutic interventions. This review focuses on the structural and cellular alterations triggered by HCM-causing mutations in troponin and tropomyosin proteins and how structural biology can aid in the discovery of new platforms for therapeutics. We highlight the importance of a better understanding of allosteric communications within these thin-filament proteins to decipher the HCM pathological state.Inherited myopathies affect both skeletal and cardiac muscle and are commonly associated with genetic dysfunctions, leading to the production of anomalous proteins. In cardiomyopathies, mutations frequently occur in sarcomeric genes, but the cause-effect scenario between genetic alterations and pathological processes remains elusive. Hypertrophic cardiomyopathy (HCM) was the first cardiac disease associated with a genetic background. Since the discovery of the first mutation in the β-myosin heavy chain, more than 1400 new mutations in 11 sarcomeric genes have been reported, awarding HCM the title of the “disease of the sarcomere.” The most common macroscopic phenotypes are left ventricle and interventricular septal thickening, but because the clinical profile of this disease is quite heterogeneous, these phenotypes are not suitable for an accurate diagnosis. The development of genomic approaches for clinical investigation allows for diagnostic progress and understanding at the molecular level. Meanwhile, the lack of accurate  MHY7 gene, R403Q)  triggers the release of Ca2+ from the sarcoplasmic reticulum to start contraction. Mutations in RyR2 lead to aberrant intracellular Ca2+ metabolism and Ca2+ overload that may have an involvement in arrhythmias  pattern Marian, . Its diaAlmost 50 years ago, Francis Crick published a noteworthy article entitled the “Central dogma of molecular biology” Crick, . At thatsource of information is important for producing the message. An analogy of the source could be a sequence of letters organized in a coherent way to form a word or sentence. (ii) The transmitter is responsible for sending this message through a channel. (iii) The channel links the transmitter to the receiver. (iv) The receiver makes the message intelligible when reaching its destination. (v) The destination is where the message arrives and can then perform its function  serves as the source of information. Equally important, the RNA molecule plays the transmitter role, allowing the correct message to reach the channel. The nuclear pore complex recognizes mature RNA molecules and sends them to the receiver, thus serving as a selective channel. Next, the complex machinery of the ribosome acts as the receiver, transforming the genetic information through the synthesis of a correct polypeptide sequence. In this analogy, additional receivers would allow proper polypeptide folding  and the incorporation of important modifications, such as phosphorylation and glycosylation, among others . Finally, the destination of this functional protein will generate the expected response inside or outside the cell. This simplistic analogy illustrates how any substitution occurring at the message level may significantly affect the correct communication and cause either a misunderstanding or a new understanding at the destination Figure .Correct folding is intimately linked to the function or, analogously, the message of a biomolecule. A remarkable study in the 1950s showed that the three-dimensional assembly of a protein is guided by its specific amino acid sequence. The physicochemical properties of specific amino acid side chains lead to a hydrophobic collapse event; thus, proper folding is expected to occur independently of biological cellular machinery  commonly occur in native globular proteins and result in structurally similar conformers with implications for molecular recognition  in which a small number of high-energy and short-lived conformers are populated  or gain-of-function (GoF) events occur. Upon mutation, LoF and GoF events are commonly observed in different proteins, including tumor suppressors and sarcomeric proteins, with implications in cancer and cardiac disorders  they lead to alterations in protein structure and function and (ii) changes frequently occur in highly conserved amino acids throughout evolution  and the process of alternative cleavage and polyadenylation (APA) of mRNAs as regulators of gene expression. For instance, downregulation of a specific miRNA locus in stressed cardiomyocytes is sufficient to attenuate the increase of cell size  and the troponin complex (Tn). Actin is a ubiquitously expressed protein and participates in various cellular events, such as motility, cytokinesis and contraction. Although some actin mutations are involved in the HCM  is the direct Ca2+ sensor in the myofilament. The conformational changes triggered by Ca2+ binding at specific cTnC sites control the allosteric signaling cascade along the entire complex. Cardiac troponin I (cTnI) performs the classical role of ATPase activity inhibition. In addition, its tridimensional structure plays an important regulatory role in protein-protein interactions. The cardiac troponin T (cTnT) is the “molecular glue” that anchors the Tn members to the thin filament, playing an important role in Ca2+ transduction structural signaling. Together, the Tn complex and Tm represent the regulatory proteins of the thin filament  containing a highly conserved amino acid sequence among TnI isoforms; (v) a switch region (cTnI147−163) that experiences a disordered-ordered transition upon binding to the N-terminal domain of cTnC-triggering contraction  an d Ramos, ; (ii) and Ramos, ; (iii) a2+ stabilize a cTnI conformation that suppresses the power stroke and thus prevents actin-myosin interactions, mostly because the cTnI inhibitory region is sitting on actin , releasing actin inhibition  and R21C cTnI mutants altered the PKA-dependent effects on weakening cTnC-cTnI interactions and accelerating myofibril relaxation  presented cardiomyocyte disarray, fibrosis, and hypercontractility with diastolic dysfunction, symptoms that align with an increased sensitivity to Ca2+ switch of the thin filament   and competitively bind Mg2+ (~103 M−1). Although the cNTnC has two Ca2+ binding sites, site I is inactive due to several loop substitutions that impair Ca2+ coordination, for example, the inclusion of hydrophobic residues instead of charged residues in the +X and +Y positions  the binding of Ca2+ at site II and (ii) the interaction between cTnC and cTnI leading to changes in the number of hydrophobic patches and to the further stabilization of the open active state. Both factors are essential for contraction.Different TnC isoforms are expressed in human cardiac/slow skeletal and fast skeletal muscle cells and are encoded by the 2+ and the cTnC cellular partner interactions. Currently, the frequency of cTnC mutations is comparable to that of other targets, such as α-Tm and actin  levels was observed in A8V heart samples. This finding agrees with recent investigations suggesting that in humans, sarcomeric mutations have decreased the levels of cTnI-P  integrate this multigene family in vertebrates, and each gene can produce different splicing isoforms in specific tissues. Of note, α-Tm is the predominant isoform expressed in the heart and comprises 284 residues. Gene shuffling leads to the expression of at least 40 Tm isoforms, and this multiplicity seems to play a pivotal role in cell maintenance  has been proposed in an attempt to explain this dynamic and allosteric actomyosin regulation. Under resting conditions and low Ca2+ levels, the regulatory site of TnC is empty, characterizing the blocked state. Additionally, the interaction between actin and TnI is stabilized, and Tm locks actin-myosin interactions. The closed state is characterized by the binding of Ca2+ to the regulatory domain of cTnC and the exposure of a hydrophobic patch that further interacts with the C-terminal domain of cTnI. Moreover, the inhibitory peptide of cTnI moves away from actin. This transition state is characterized by the partial exposure of the myosin binding sites triggered by an azimuthal motion of Tm over the actin filament, generating weakly bound cross-bridges  and skinned fibers (higher Ca2+ sensitivity for the mutant). Interestingly, an evaluation of biopsies from two patients carrying the HCM-related α-Tm mutant D175N revealed the in vivo incorporation of the mutation within the vastus lateralis muscle, resulting in an altered contractile function , but mutant homodimers have a slightly slower affinity compared to wt  but, surprisingly, no changes in the myofilament Ca2+ sensitivity or the response to β-adrenergic challenges  flanked by packed domains. The complexity of these multi-domain architectures was in the past attributed only to their folded modules, but recent investigations and emerging techniques in structural biology have shown the key participation of IDRs as dynamic elements triggering signaling hierarchy and tuning protein functionalities. This new “2+ to the C subunit of the troponin complex. The linker connecting the TnC structural and regulatory domains was shown to communicate both regions in a synergistic way  leads to an increased affinity of the Ca2+-binding site II  is not observed in current crystal structures (PDB codes 1J1E and 4Y99), limiting the structural and functional characterization of this inhibitory segment for peptide assays  relaxation dispersion experiments are sensitive to millisecond motions  was validated as a tool for producing isotopically labeled Tn peptides for NMR structural and drug-screening tests , has the potential to reproduce physiological and pathological phenotypes Adult cardiomyocytes are differentiated and complex cells playing an unambiguous role in cardiac excitation-contraction coupling. This is achieved through a fine-tuned communication signaling of hundreds of molecules organized in weakly and tightly bound molecular assemblies, including cytosolic and transmembrane proteins, ion channels, transcriptional factors, adaptors, and second messengers Figure . UnfortuRegardless of the sarcomeric repertoire of mutations causing HCM and heart tolerability to mechanical stretch upon heartbeat, the pathways discriminating physiological and pathological mechanosensitivities remain scarce. The correct tuning of biomechanical stress and how molecules sense and trigger hypertrophy is a matter of debate, but proteins located at the Z disc and at the costamere complex may play a role in this mechanotransduction process. The Z disc is responsible for transferring the tension between the sarcomeres through the interaction between titin and α-actinin . Thus, NFAT translocates to the nucleus and reactivates the fetal gene program, which includes the expression of the brain natriuretic peptide (BNP), α-skeletal actin and β-myosin heavy chain (β-MHC) . These receptors are activated by factors released upon increased pressure or mechanical stretch, such as angiotensin II and endothelin (ET1), and play a role in hypertrophic responses. Additionally, GPCR-dependent PLCβ activation can lead to CaThe production and deposit of collagen type I and III by fibroblasts are other alterations observed in early cardiomyopathy characterizing fibrosis. Increased deposits may impair the excitation-contraction coupling, leading to severe changes in heart contractility (Menon et al., Several lines of evidence also link the activation of apoptotic signaling in hypertrophic hearts. An investigation of explanted failing hearts from transplanted patients with cardiomyopathy revealed increased levels of cytochrome C associated with the activation of the pro-apoptotic cysteine-aspartic acid protease 3, caspase-3 (Narula et al., sine qua non condition for better understanding the thin/thick filament interactions and ultimately the muscle contraction phenomena under normal and pathological conditions. The impact of mutations and post-translational modifications in sarcomeric proteins and their effects on the generation of non-adaptive hypertrophy are clarified from a series of well-designed studies. Because this review was written by structural biologists, the contribution that should come from this paper is the provision of a point-by-point investigation of the short- and long-term allosteric changes in different pathogenic mutations on isolated proteins and within the context of their biological partners. Because HCM presents a heterogeneous profile of genetic inherence, deciphering the altered motions and dynamic pathways through personalized mutagenic studies will certainly provide insight for new drugable sites that would have the potential for future therapeutics to hopefully ameliorate the hypertrophic phenotype from a broader range of pathogenic mutants.The combination of multidisciplinary expertise in structural biology, biochemistry, physiology, and cellular biology represent a All authors listed, have made substantial, direct and intellectual contribution to the work, and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Zinc is a metal ion that is essential for growth and development, immunity, and metabolism, and therefore vital for life. Recent studies have highlighted zinc’s dynamic role as an insulin mimetic and a cellular second messenger that controls many processes associated with insulin signaling and other downstream pathways that are amendable to glycemic control.Mechanisms that contribute to the decompartmentalization of zinc and dysfunctional zinc transporter mechanisms, including zinc signaling are associated with metabolic disease, including type 2 diabetes. The actions of the proteins involved in the uptake, storage, compartmentalization and distribution of zinc in cells is under intense investigation. Of these, emerging research has highlighted a role for several zinc transporters in the initiation of zinc signaling events in cells that lead to metabolic processes associated with maintaining insulin sensitivity and thus glycemic homeostasis.This raises the possibility that zinc transporters could provide novel utility to be targeted experimentally and in a clinical setting to treat patients with insulin resistance and thus introduce a new class of drug target with utility for diabetes pharmacotherapy. Insulin resistance (IR) is a common pathophysiological condition in which patients present with reduced insulin sensitivity and thus glucose intolerance, particularly in liver, adipose tissue and skeletal muscle . This haRecently, research on metabolic processes associated with IR and T2DM has revealed an exciting role for the biochemical and physiological role of zinc and the proteins that transport zinc in cells in diseases associated with abnormal cellular signaling . AccordiZinc is ubiquitous in physiological systems, albeit, within tightly controlled parameters, and therefore suggests that atypical levels are likely to have significant biological and clinical effects on disease processes. Knowing how zinc transporter proteins and the storage of zinc in cells are involved in metabolic processes implicated in IR for example, may present opportunities to develop novel drugs targeting these transporters to prevent or treat IR and T2DM disease progression.Type 2 diabetes mellitus (T2DM) is devastating disorder characterised by hyperinsulinemia, hyperglycaemia, compromised energy metabolism and expenditure, and the progression of chronic illness and disease. T2DM is high complex involving both genetic predisposition and environmental factors. A major factor involved in a person’s susceptibility to T2DM can be linked through family history of diabetes. For example, Pacific Islander peoples are a unique population with especially high rates of T2DM . The envThe development of T2DM is preceded by IR, a disorder associated with hyperinsulinemia, glucose intolerance and dysfunctional energy metabolism . A leadiThere are a range of medicines available to manage and treat IR and T2DM, however, side effects of these drugs have always been a foremost challenge in relation to the goal of pharmacotherapy  Table 11. Yet, tZinc is an essential trace element that is found in all parts of the body including the fluids and secretions, tissues and organs. Zinc is one of the most abundant trace metals (next to iron) in the human body, containing approximately 2–4 g . The conTwo families of zinc transporter proteins and zinc buffering proteins play a critical role in the influx, efflux, buffering and compartmentalization of zinc. These are the zinc transporters (ZnT proteins and Zrt/Irt-like ZIP proteins), and the intracellular zinc-binding metallothionein (MT) proteins . MT protSaccharomyces cerevisiae (designated ZRT1) based on an amino acid sequence similarity to that of Irt1p, an Fe(II) regulated transporter from Arabidopsis thaliana [The original member of the ZIP family of zinc transporters was identified in thaliana , 21. Conn, n = 3–5) situated between TMD 3 and TMD 4 that is suggested to serve as a zinc-binding site  structure and this is orientated with the N- and C-terminal facing the extracytosolic region. Many of these members have a predicted long histidine-rich loop region  early zinc signaling (EZS) and, 2) late zinc signaling (LZS). EZS, is a process that is independent of gene transcription and results in rapid changes in intracellular levels of zinc that occurs in minutes (the ‘zinc wave’ response) through the triggered release of zinc from subcellular organelles into the cytosol . This phThe defining role of zinc as a signaling molecule was shown in early studies in rat adipocytes where zinc could stimulate lipogenesis, independent, and additive to that of insulin . SimilarThe mechanisms of the insulin-mimetic action of zinc have been delineated in several studies however it is still unclear how these processes occur. One well-established mechanism of zinc action on cellular signaling events occurs through the inhibition of protein tyrosine phosphatase 1B (PTP1B). PTP1B functions as a negative regulator of insulin and leptin signaling transduction pathways . Thus, tGiven the well-established role of zinc transporters in mediating the critical control of zinc homeostasis in cells, it will be important to further delineate their function in cell signaling events in the context of metabolic control. Currently, information is limited to what role the zinc transporters might play in cell signaling events in the context of insulin resistance. Therefore, extrapolation of studies from other cellular systems or disease states that have identified zinc transporters and thus zinc flux in facilitating cell signaling events might prove useful.Studies accessing the role of zinc transporters in cellular signaling found that the zinc transporters ZnT5 and ZnT7 are responsible for loading zinc to alkaline phosphatases (ALPs) in the biosynthetic-secretory pathway in chicken B lymphocyte-derived cells . These aXenopus oocytes and cultured mammalian cells [In other studies, ZnT1 can regulate Raf-1 enzymatic activity in an cells . Raf-1 pan cells . ZnT1 waan cells  suggestic-fos activity was significantly reduced. PTHrP stimulates the phosphorylation of cAMP response element-binding protein (CREB) which in turn induces the transcription of c-fos [Of the ZIP transporters, ZIP14 is important in G-protein coupled receptor (GPCR)-mediated signaling through the maintenance of intracellular cAMP levels via suppression of phosphodiesterase activity . In thesof c-fos .Studies in MCF breast cancer cell lines have identified that ZIP7 is essential in the redistribution of zinc from intracellular stores to the cytoplasm and subsequent zinc-induced inhibition of phosphatases . MoreoveZIP7 is essential for the proliferation of intestinal epithelial cells and mice lacking this transporter in intestinal epithelium have massive apoptosis of transit-amplifying cells due to increased endoplasmic reticulum stress (ER) . SimilarPreviously we have identified a role for ZIP7 in glycemic control in skeletal muscle . SkeletaRecent studies have shown a relationship with ZIP13 and beige adipocyte biogenesis and thermogenesis . In primZIP14 has been identified as a critical route for non-transferrin bound iron (NTBI) uptake into liver and pancreatic acinar cells and is essential for the development of liver iron overload in hemochromatosis . DiabeteZinc plays a major role in many aspects of cell signaling events in several physiological and pathophysiological processes. While it is well established that at least one zinc transporter (ZnT8) is critical for the compartmentalization, structure and secretion of insulin in beta cells of the pancreas , there iZinc is critical for the physiological role of insulin in the form of storage in the secretory granules of the pancreas as an inactive zinc-insulin hexamer , 62. WheGenome-wide association studies (GWAS) have discovered that a nonsynonymous single nucleotide polymorphism (SNP) in ZnT8 rs13266634) encodes a C → T base substitution resulting in a change in the coded protein (p.Arg325Trp) and the production of two protein variants R and W of which the C allele (R variant) is associated with susceptibility to type 2 diabetes . The fre634 encodAlthough GWAS studies have been successful in identifying variant ZnT8 alleles, it does not necessarily imply that the risk allele has a direct pathophysiological effect on beta-cell insulin secretion. A Meta-Analysis of Glucose and Insulin-related traits Consortium (MAGIC) was recently formed to conduct large-scale meta-analyses of genome-wide data for continuous diabetes-related traits in non-diabetics . Meta-anMore recent studies that sequenced approximately 150,000 individuals across five ancestry groups reported twelve rare protein-truncating mutations in ZnT8 which together explain a 65% reduced risk of developing type 2 diabetes . It was Recently, studies aimed to delineate the effect of dietary factor interactions with ZnT8 polymorphism (rs13266634) and the risk of developing metabolic syndrome found a significant interaction among omega-3 fatty acid consumption and ZnT8 in the context of metabolic syndrome, dyslipidemia, and abdominal obesity . ParticiStudies to investigate the role of ZnT8 and glucose homeostasis has been established with ZnT8 null mouse models with global deletion of ZnT8 or pancreatic beta-cell specific ZnT8 deletion . Most moGiven the pancreatic tissue-specificity of ZnT8 there is potential for this transporter to be amendable to therapy in the treatment of diabetes. However, targeting ZnT8 in the treatment of diabetes could prove to be highly complex. Although results from ZnT8 null mice suggest that increasing ZnT8 could improve insulin secretion and glycemic control, loss of function mutations in ZnT8 suggest a protective role for this transporter in type 2 diabetes. While ZnT8 plays a critical role in insulin physiology, and over the last decade or so this transporter has been given much attention for its role in diabetes, other zinc transporters have not had the same focused attention until recently. Apart from the many studies on zinc signaling in cells and the insulin-mimetic action of zinc, it is unclear which zinc transporters are involved in initiating these signaling processes. Clues from studies on the role of ZIP7 as the “gate-keeper” of zinc release from the Golgi apparatus  and subsZinc is an essential metal ion that is ubiquitous in many metabolic and physiological processes. The emerging role of zinc as an insulin mimetic in maintaining cellular function suggests that atypical levels, and aberrant compartmentalization, transport and storage of zinc will have biological effects that could be amendable to clinical intervention. Although current understandings on the role of zinc transporters in insulin resistance is not available, and this knowledge is only just emerging in type 2 diabetes, it is clear from studies on ZnT8 that this family of transporters has utility for the development of novel diabetic therapies. While ZnT8 plays a significant role in insulin biology and therefore represents an attractive target for diabetes therapy, the other members of the zinc transporter family in diabetes are less defined. However, we can speculate from the information presented in this review that the other transporters are involved in processes that facilitate insulin signaling and glycemic control and therefore could offer exciting new targets that are amendable to therapeutic intervention in the treatment of diseases associated with insulin resistance and type 2 diabetes.


High-tension electricity can cause devastating injuries that may result in abdominal wall loss, visceral damage, and sometimes major threat to life. The visceral organ may be exposed after debridement and require flap cover, but the tensile strength of abdominal wall may be lack even if flap transplanted.From April 2007 through May 2015, 5 patients with severe abdominal electrical injury were treated at our hospital. Exploratory laparotomy was performed based on their clinical manifestations and debridement findings of abdominal wall at early stage, and decision regarding technique for reconstruction of abdominal wall was based on an assessment of the location and extent of the defect. Medical records were reviewed for these data.Clinical evaluation and debridement findings of the abdomen revealed 4 patients with suspicious visceral damage. Laparotomy was performed in 4 cases, and revealed obvious lesion in 3 cases, including segmental necrosis of small intestine, partial necrosis of diaphragm, left liver and gastric wall, and greater omentum. Five patients underwent abdominal wall reconstruction using island retrograde latissimus dorsi myocutaneous flap or free/island composite anterolateral thigh myocutaneous flap. All flaps survived, abdominal bulging occurred in 3 cases after follow-up of 12 to 36 months.The clinical manifestations and wound features of abdomen collectively suggest a possible requirement of laparotomy for severe abdominal electrical burns. Retrograde latissimus dorsi myocutaneous flap or composite anterolateral thigh myocutaneous flap is an effective option for reconstruction of abdominal wall loss, the long-term complication of abdominal bulging, however, remains a significant clinical challenge. Exploratory laparotomy can find ischemic or necrotic tissue, perforate hollow organ, and other damage, but sometimes no obvious lesion can be found after laparotomy, the indication of exploratory laparotomy is still deserved to be investigated.Electrical injuries on abdominal wall are uncommon, but associated with high mortality, especially delayed gastrointestinal perforation and peritonitis complicated after the visceral organ injury.–7 A series of free or pedicled anterolateral thigh (ALT) and/or tensor fasciae latae (TFL) flaps can be chosen for abdominal wall reconstruction depending on the size and location of the defect and the length of the vascular pedicle.–17 The benefits of flap cover in the acute stage are early wound coverage, preservation of vital viscera and digestive function, reduced invasive infection, and hospital stay. But the tensile strength of abdominal wall may be lack even if flap transplanted, a flap with strong, vascularized fascia and/or innervated muscle can enhance the tensile strength of abdominal wall to avoid abdominal hernia formation .,15,19–20 Therefore, the reconstruction of large defects on the abdominal wall after electrical injury can be challenging. The purpose of this article is to retrospectively analyze the characteristics of abdominal electrical injury with visceral damage and to evaluate the outcome of early flap repair of large defects on the abdominal wall after electrical burns.The basic principle of electrical burns, especially visceral organ exposed after debridement, is to repair the defect with a vascularized flap.22.12 (mean 494.4 cm2). After admitted, meticulous symptom inquiry and detailed physical examination, if needed, serial accessory examination such as abdominocentesis, KUB, ultrasonographic examination, and so on were performed. Besides, 4000 to 6000 mL fluid was administrated for fluid replacement to maintain a clear hourly urine output to 50 to 75 mL/h.Five patients suffering from a high-voltage electrical injury to extensive abdominal wall were treated at our academic hospital from April 2007 to May 2015. All of the patients were male with an average age of 46 years . The induced voltage of these patients was 1000 to 100,000 V and they were evaluated on admission between 16 and 24 hours (mean 19 hours) after burn injury. None of the patients had any chronic illnesses, such as diabetes or cardiovascular insufficiency, nor did they have cardiogenic or hypovolemic shock symptoms or experience other major trauma, such as an intracranial injury or a bone fracture. The majority of patients’ limbs were injured or even damaged, and the calculated total burned body surface area of these patients ranged from 3% to 19% (mean 11.0%). The size of abdominal wall wound ranged from 360 to 700 cmConsents to conduct and report of this study were obtained from the Ethics Committee of Xiangya Hospital, Central South University. Written informed consent for publication of this case report and any accompanying images was obtained from the patient.2.2Debridement was performed within 1 to 3 days after injured. Eschar and obvious devitalized muscle or other tissues were removed, the appearance of viable abdominal muscle, posterior rectus sheath, or peritoneum were noted and left as much as possible during debridement. All patients were debrided for one time. According to the clinical manifestations mentioned above and debridement findings at early stage, laparotomy operations were performed for those under suspicion with internal organs injury. During laparotomy intestinal resection and anastomosis, or resection of necrotic tissues  and repair of visceral organ were performed, and the peritoneal cavity was closed with peritoneum or greater omentum.2.3After debridement and laparotomy, one-stage reconstruction followed. The defect of abdominal wall was repaired with island retrograde latissimus dorsi (LD) musculocutaneous flap, free, or island composite ALT musculocutaneous flap. The selection of flap for abdominal wall reconstruction was determined by the location and size of the defect. During harvest of the former perfused by lumbar artery and perforator intercostal vessels, a major perforator from the 10th intercostal vessels was secured. When harvesting of the latter perfused by the lateral circumflex system of femoral vessels, vastus lateralis, or rectus femoris and the femoral nerve of these muscle were carried if necessary. The deep inferior epigastric vessels were used as the recipients for end-to-end anastomoses during free-flap transplantation. To strengthen the abdominal wall, the carried vastus lateralis or rectus femoris was sutured with remnant rim of abdominal muscles including the remaining rectus abdominis using numerous strong prolene sutures, the fusion of fascia lata was fixed to the external oblique aponeurosis under suitable tension, and the femoral nerve of vastus lateralis muscle was coapted with the intercostal nerve when free transplanted. The immediate and long-term outcomes were assessed regarding the complications and sustainable strength of the abdominal wall. The patient characteristics are shown in Table 33.1On physical examination, an abdominal and lumber burn wound of about 20 cm × 15 cm was detected, and we found a direct deep wound to the liver and stomach surface through the necrotic wound of the left 9th and 10th ribs, a 2 cm × 3 cm area of necrosis of the diaphragm, marginal necrosis of left liver and a necrotic pitting in the anterior wall of stomach with no obvious perforation during debridement. After resection and repair of injured diaphragm and gastric wall, we used retrograde LD musculocutaneous flap to cover the defect  presented with pitting and contracture of the intestine, and edema was noted in the remaining three sites. After the repair of the 3 edematous lesions, the necrotic intestines were resected and followed by enteroanastomosis. Right island composite ALT musculocutaneous flap was designed to close the abdominal defect. The abdominal wall was strengthened by suturing up the vastus lateralis with the rectus abdominis, and the fusion of fascia lata with the external oblique aponeurosis. The residual defects were covered by split-thickness skin grafts. On day 8 in hospital, the patient underwent amputation of the left forearm and the right arm, and debridement of the left thigh. On day 26 in hospital, free LD musculocutaneous flap was designed to cover the defect with bone exposure in the left thigh. Thirty-six months after repair, the reconstructed abdominal wall was bulging without herniation of intraperitoneal organs  or autologous material (tissue flaps). and the significant risk of long-term complication should be advised to patient. In addition, possible donor site morbidity should also be considered. As Kuo et al[18] reported quadriceps femoris muscle contraction forces of donor thigh was reduced, on average, by 30% after harvest of an ALT flap combined with vascular fascia lata (without fascia lata muscle) and a cuff of vastus lateralis muscle in spite of the fact that no difficulties in daily ambulating occurred. In consideration of the long-term outcomes of cases 3, 4, and 5, the majority of abdominal bulging could be considered to be dependent on the lack of muscle and fascia structure with tensile strength, some of which is associated with the swollen wound and disability obesity after electrical burns. Therefore, we suggest that some adjacent muscle may be carried with the ALT flap used to repair the defect on the middle or lower part of abdominal wall.Long-term follow-up showed 3 cases with abdominal bulging; however, surgical revision was not necessary because no functional discomfort occurred. The similar outcome of abdominal bulging has been reported by Bodin,,19,34 but the recipient nerve is very difficult to find and not suit for anastomose with the nerve of flap owing to length and orientation in some cases. We used an innervated free ALT musculocutaneous flap in case 2, and pedicled innervated vastus lateralis or rectus femoris muscle in case 3 and 5, respectively, the result of follow-up is passable, especially distinct bulging appeared only in the left wound that repaired without innervated muscles in case 5, but the morbidity of donor-site is more obvious of course, and more extensive investigations with comparative studies are needed. By this token, the perfect reconstruction of abdominal wall loss after electrical injury still remains a significant clinical challenge.Previous publications about the reconstruction of abdominal wall defect with an innervated free LD or ALT musculocutaneous flap, the thoracodorsal nerve or femoral nerve of the vastus lateralis muscle was coapted with the intercostal nerve,


Traditional genomic prediction models using multiple regression on single nucleotide polymorphisms (SNPs) genotypes exploit associations between genotypes of quantitative trait loci (QTL) and SNPs, which can be created by historical linkage disequilibrium (LD), recent co-segregation (CS) and pedigree relationships. Results from field data analyses show that prediction accuracy is usually much higher for individuals that are close relatives of the training population than for distantly related individuals. A possible reason is that historical LD between QTL and SNPs is weak and, for close relatives, prediction accuracy of SNP models is mainly contributed by pedigree relationships and CS. Information from pedigree relationships decreases fast over generations and only contributes to within-family prediction. Information from CS is affected by family structures and effective population size, and can have a substantial contribution to prediction accuracy when modeled explicitly.In this study, a method to explicitly model CS was developed by following the transmission of putative QTL alleles using allele origins at SNPs. Bayesian hierarchical models that combine information from LD and CS (LD-CS model) were developed for genomic prediction in pedigree populations. Contributions of LD and CS information to prediction accuracy across families and generations without retraining were investigated in simulated half-sib datasets and deep pedigrees with different recent effective population sizes, respectively. Results from half-sib datasets showed that when historical LD between QTL and SNPs is low, accuracy of the LD model decreased when the training data size is increased by adding independent sire families, but accuracies from the CS and LD-CS models increased and plateaued rapidly. Results from deep pedigree datasets show that the LD model had high accuracy across generations only when historical LD between QTL and SNPs was high. Modeling CS explicitly resulted in higher accuracy than the LD model across generations when the mating design generated many close relatives.Our results suggest that modeling CS explicitly improves accuracy of genomic prediction when historical LD between QTL and SNPs is low. Modeling both LD and CS explicitly is expected to improve accuracy when recent effective population size is small, or when the training data include many independent families.The online version of this article (doi:10.1186/s12711-016-0255-4) contains supplementary material, which is available to authorized users. The feasibility of obtaining genotypes of dense single nucleotide polymorphisms (SNPs) with genome-wide coverage has improved accuracy of estimated breeding values by genomic prediction –6. To daCo-segregation is an important source of information that contributes to accuracy of genomic prediction , 10. AllIn analyses of field datasets using SNP models, high accuracy of genomic prediction has been mainly observed among close relatives , 15, 16,Simulation studies have shown that both LD and CS information contribute to prediction accuracy of the genomic best linear unbiased prediction (GBLUP) model . InformaAlthough LD between SNPs has been shown to be sizable in livestock populations , 22–24, Explicit modeling of CS information for genomic prediction was proposed and developed by Luan et al. . In LuanPersistence of prediction accuracy across validation generations without retraining (long-term accuracy) is an important criterion to evaluate contributions of LD and CS information to prediction accuracy. Habier et al.  showed tRecent effective population size (The objectives of this study were (1) to develop a Bayesian statistical method to model CS explicitly by following the transmission of putative QTL alleles of pedigree founders, (2) to investigate contributions of LD and CS information to the accuracy of genomic prediction across unrelated families and validation generations without re-training, and (3) to investigate the effects of historical LD, recent Definitions of LD, CS, and pedigree relationship are presented for a population with known pedigree information following Habier et al. . LD is dn training individuals, m SNPs of each training individual, m SNPs, and Following Meuwissen et al. , the staIn the LD model , QTL effn training individuals, jth putative QTL, with The CS model is given bymentclass2pt{minimThe model that fits both LD and CS (LD-CS model) includes the LD and CS terms from models  and 2),,2),3\docCo-segregation of alleles at two loci means that these alleles share identical grand-parental allele origins, i.e. they both originate from the same chromosome of a parent. The indicator of parental allele origin at one locus is a Bernoulli variable. In this study, the allele origin indicator equals 0 if it originates from its grand-maternal allele, and 1 if it originates from its grand-paternal allele. When allele origins of parents and offspring at a SNP are known, the probability that the allele origin of a putative QTL linked to the SNP is grand-paternal  can be calculated using recombination rates between QTL and SNPs, which is termed the probability of descent of the QTL allele (PDQ). Suppose that allele origins are known for an individual’s maternal alleles at two SNPs unctions .In most cases, positions of QTL are not known, and genome-wide CS information is modeled for putative QTL within each non-overlapping genomic window of a certain length. In this study, putative QTL were fitted at genome windows of lengths 0.5, 1 and 2 cM, respectively, to study the impact of window length on the accuracy of the CS and LD-CS models. Within each genomic window, the putative QTL is assumed to be located at the midpoint of SNPs i  of an individual is the summation of the value of its maternal and paternal alleles  [In pedigree 2, all 300 sires and 300 dams from generation 2 were used as parents. Each sire was mated with one dam, producing one male and one female progeny. Pedigree 2 represents an outbred population where all individuals survived, but each individual had a relatively limited contribution to future generations. Since each individual contributes an equal number of gametes to the next generation, and the variance of family sizes is zero, the igree 2) .In pedigree 3, five sires and 70 dams were randomly selected in each generation starting from the second generation. One sire was mated with 50 dams, each dam producing five male and five female progeny, representing an influential sire family. Each of the other four sires was mated with five dams, each dam producing two male and three female progeny, representing four small sire families. Pedigree 3 represents an unbalanced nested design, where the genetics of one individual dominates future generations. The The first five pedigree generations, with size 2455 (pedigree 1 and 2) and 2475 (pedigree 3), were used for training. Each of the following eight generations, with size 600, were used for validation. Prediction accuracy was calculated as the correlation between GEBV and TBV in each validation generation.In all simulated datasets, allele origins were assumed known without error at all SNPs. Putative QTL were fitted within each 1-cM genome and PDQ were calculated using the simulated true allele origins. In addition, for pedigree 1, allele origins at all SNPs were either assumed known without error, or imputed using the LDMIP software, with default parameter settings . PutativIn the Common QTL scenario with high historical LD, prediction accuracy of the LD model increased from less than 0.2 with one half-sib family and quickly plateaued around 0.8 when the number of half-sib families exceeded 50, which corresponds to a training size of 500 Fig. . AccuracIn the Rare QTL scenario with high historical LD between SNPs, the actual level of historical LD between QTL and SNPs was low due to all QTL having much lower MAF than SNPs. Accuracy of the LD model increased with training size from 0.1 to about 0.45, which was much lower than accuracy with the Common QTL scenario  to develop a Bayesian statistical method to model CS information explicitly, (2) to study contributions of LD and CS information to accuracy of genomic prediction across unrelated families and validation generations without re-training, and (3) to study the effects of historical LD, recent In this study, a new method that explicitly models CS information was developed for genomic prediction in pedigree populations. This method models transmission of putative QTL alleles within consecutive non-overlapping genomic windows of sufficiently small length (1 cM in this study), such that the recombination rate is so small that the alleles at all polymorphic loci within the window are expected to co-segregate for several generations. Co-segregation of QTL alleles was modeled using parental allele origins at SNPs that cover the genomic window, which are independent of the level of LD between QTL and SNPs. This method is applicable to any type of pedigree population provided that allele origins are available (or can be imputed) from founders to offspring. The method of modeling CS at putative QTL using allele origins at observable SNPs is similar to the method developed by Fernando and Grossman , but hasn) times the number of founder QTL alleles n because it requires the inverse of a dense matrix, The CS model  developen et al. , the CS entclass1pt{minimaentclass1pt{minimaentclass1pt{miniman et al. , computaIt is generally accepted that LD between QTL and SNPs, co-segregation of QTL with SNP alleles, and pedigree relationships at QTL captured by SNPs are the three main sources of information that contribute to the accuracy of genomic prediction , 17, 34.In the simulated datasets with different numbers of unrelated half-sib families, both LD and CS information contributed to the accuracy of genomic prediction. Accuracy of the LD model relies on the level of LD between QTL and SNPs in the base population. Accuracy of the CS model relies on accurate estimation of the value of founder QTL alleles that are transmitted to half-sib offspring within the same family. Accuracy of the LD model increased rapidly with increasing training size when LD between QTL and SNPs was high, because high LD is conserved across families and increasing training size brings in more data to improve estimation of SNP effects. However, when historical LD is low or zero, accuracy of the LD model mainly comes from capturing CS information, which only exists within the same half-sib family. Consequently, with more unrelated half-sib families included in the training population, accuracy of the LD model decreased and became lower than accuracy of the CS model. This is because only half-sibs from the same family contribute to prediction accuracy. In the CS model, the value of founder QTL alleles is estimated using only information from the same half-sib family, while the LD model estimates SNP effects by pooling CS information across all families, which is erroneous because linkage phase and LD is highly variable across a large number of unrelated families.Three mating designs were simulated that differed in the number of parents per generation and the number of progeny per mating. Pedigrees 1 and 3 resemble the breeding program where a few sires are selected and intensively used for breeding in each generation. CS information made significant contributions to prediction accuracy in pedigrees 1 and 3, because a limited number of sire alleles segregated among a large number of their progeny, and the value of the sires’ alleles can be estimated accurately based on the amount of data available. Pedigree 1 is a balanced nested design with identical family sizes, which is similar to the structure of nucleus herds in swine  or poultThe difference between the three mating designs can be quantified by recent The contribution of LD information to prediction accuracy should not depend on the mating design because high historical LD between QTL and SNPs is mostly between closely linked loci and hardly erodes within several recent generations. However, because the LD model also implicitly captures information from CS and pedigree relationships , accuracIn general, when historical LD is high between QTL and SNPs, long-term accuracy is mostly contributed by LD information and CS information has little contribution regardless of recent LD quantifies the correlation between allele states at QTL and SNPs. The LD model captures this correlation using multiple regression on SNP genotypes. Strong correlations can only exist when QTL and SNPs have similar MAF, as in the Common QTL scenario. Correlations are expected to be low when most QTL have low MAF, while SNPs have moderate MAF, as in the Rare QTL scenario. In the simulated datasets, the correlation between allele states exists in the form of historical LD and recent CS. Historical LD between closely linked loci is hardly eroded by recombination. The correlation generated by CS can exist between loci over long chromosome regions, which erodes fast with recombination. Both forms of correlation can be captured by the LD model, however, the LD model has persistent long-term accuracy only when historical LD between QTL and SNPs is high, which requires similar MAF between QTL and SNPs. When historical LD is low due to most QTL having low MAF, prediction accuracy of the LD model mainly comes from implicitly capturing CS information, which decreases rapidly across validation generations because CS information across long chromosome regions erodes fast with recombination. Similar results have been observed by Habier et al.  for the CS follows the transmission of QTL alleles among related individuals, which is independent of historical LD. As a result, prediction accuracy from the CS model is not affected by the level of historical LD. Accuracy due to CS information depends on (1) the length of the founder haplotypes that are used to follow transmission of putative QTL alleles, which determines the rate of erosion of CS due to recombination; and (2) the accuracy with which the value of founder alleles can be estimated, which depends on the amount of phenotype data for the progeny that inherited the same founder haplotype. The CS model that used haplotypes of length 1 cM is expected to have only a few recombinations within haplotypes, and therefore CS information contributes to long-term accuracy provided that the value of founder QTL alleles can be estimated correctly with sufficient data.t prior distributions are given to t prior distribution with degrees of freedom t distribution elsewhere. BayesB results in much heavier shrinkage towards zero than BayesA, and consequently the effective number of loci fitted in the model is larger in BayesA than in BayesB [Prior distributions in BayesA and BayesB  were use1+β2νS2) . The pos1+β2νS2) . This meThe effect of using alternate prior distributions on the LD model is twofold. When historical LD is high between QTL and SNPs, high prediction accuracy is usually achieved by the LD model, with nearly unbiased estimates of large SNP effects, while effectively shrinking small SNP effects towards zero. In the simulated datasets of this study, BayesB resulted in higher accuracy than BayesA because the number of QTL was much smaller than the number of SNPs. When historical LD is low, prediction accuracy mainly comes from implicitly capturing CS information, which depends on the effective number of SNPs fitted in the model. Then, BayesA results in higher accuracy than BayesB due to fitting relatively more SNPs, which can capture more CS information than BayesB .The effect of prior distributions on the CS model does not depend on historical LD, but does depend on the MAF of QTL. When the MAF of QTL is high, BayesA tends to result in higher accuracy than BayesB because of fitting more founder alleles that co-segregate with common QTL alleles. When the MAF of QTL is low, BayesB tends to result in higher accuracy than BayesA, because only a small proportion of founder alleles carry QTL and their values can be estimated more accurately by variable selection of BayesB.In real livestock populations, persistently high accuracy across validation generations using the LD model has rarely been observed , 16–18, There are several computational issues in implementing the CS or LD-CS models  in fieldfounders , 39, 40.In this study, a new method that explicitly models co-segregation information was developed for genomic prediction of breeding values. Breeding values in this CS model were modeled as the sum of independent values of putative QTL alleles among pedigree founders, which were traced down in the pedigree using SNP haplotypes. When the training size was increased by adding unrelated half-sib families, accuracy of the CS model increased and plateaued, but accuracy of the LD model that fits SNP genotypes dropped when historical LD between QTL and SNPs was low. Modeling both LD and CS information improved prediction accuracy compared to modeling either LD or CS, especially when historical LD was low and recent CS information contributed substantially to prediction accuracy among families, which is probably the case for recent genomic evaluation in most livestock populations. The effects of recent