GASIDN: identification of sub-Golgi proteins with multi-scale feature fusion.

The Golgi apparatus is a crucial component of the inner membrane system in eukaryotic cells, playing a central role in protein biosynthesis. Dysfunction of the Golgi apparatus has been linked to neurodegenerative diseases. Accurate identification of sub-Golgi protein types is therefore essential for developing effective treatments for such diseases. Due to the expensive and time-consuming nature of experimental methods for identifying sub-Golgi protein types, various computational methods have been developed as identification tools. However, the majority of these methods rely solely on neighboring features in the protein sequence and neglect the crucial spatial structure information of the protein.To discover alternative methods for accurately identifying sub-Golgi proteins, we have developed a model called GASIDN. The GASIDN model extracts multi-dimension features by utilizing a 1D convolution module on protein sequences and a graph learning module on contact maps constructed from AlphaFold2.The model utilizes the deep representation learning model SeqVec to initialize protein sequences. GASIDN achieved accuracy values of 98.4% and 96.4% in independent testing and ten-fold cross-validation, respectively, outperforming the majority of previous predictors. To the best of our knowledge, this is the first method that utilizes multi-scale feature fusion to identify and locate sub-Golgi proteins. In order to assess the generalizability and scalability of our model, we conducted experiments to apply it in the identification of proteins from other organelles, including plant vacuoles and peroxisomes. The results obtained from these experiments demonstrated promising outcomes, indicating the effectiveness and versatility of our model. The source code and datasets can be accessed at https://github.com/SJNNNN/GASIDN .

Identification of plant vacuole proteins by using graph neural network and contact maps.

Plant vacuoles are essential organelles in the growth and development of plants, and accurate identification of their proteins is crucial for understanding their biological properties. In this study, we developed a novel model called GraphIdn for the identification of plant vacuole proteins. The model uses SeqVec, a deep representation learning model, to initialize the amino acid sequence. We utilized the AlphaFold2 algorithm to obtain the structural information of corresponding plant vacuole proteins, and then fed the calculated contact maps into a graph convolutional neural network. GraphIdn achieved accuracy values of 88.51% and 89.93% in independent testing and fivefold cross-validation, respectively, outperforming previous state-of-the-art predictors. As far as we know, this is the first model to use predicted protein topology structure graphs to identify plant vacuole proteins. Furthermore, we assessed the effectiveness and generalization capability of our GraphIdn model by applying it to identify and locate peroxisomal proteins, which yielded promising outcomes. The source code and datasets can be accessed at https://github.com/SJNNNN/GraphIdn .

Tip60/KAT5 Histone Acetyltransferase Is Required for Maintenance and Neurogenesis of Embryonic Neural Stem Cells.

Epigenetic regulation via epigenetic factors in collaboration with tissue-specific transcription factors is curtail for establishing functional organ systems during development. Brain development is tightly regulated by epigenetic factors, which are coordinately activated or inactivated during processes, and their dysregulation is linked to brain abnormalities and intellectual disability. However, the precise mechanism of epigenetic regulation in brain development and neurogenesis remains largely unknown. Here, we show that Tip60/KAT5 deletion in neural stem/progenitor cells (NSCs) in mice results in multiple abnormalities of brain development. Tip60-deficient embryonic brain led to microcephaly, and proliferating cells in the developing brain were reduced by Tip60 deficiency. In addition, neural differentiation and neuronal migration were severely affected in Tip60-deficient brains. Following neurogenesis in developing brains, gliogenesis started from the earlier stage of development in Tip60-deficient brains, indicating that Tip60 is involved in switching from neurogenesis to gliogenesis during brain development. It was also confirmed in vitro that poor neurosphere formation, proliferation defects, neural differentiation defects, and accelerated astrocytic differentiation in mutant NSCs are derived from Tip60-deficient embryonic brains. This study uncovers the critical role of Tip60 in brain development and NSC maintenance and function in vivo and in vitro.

TSNAXIP1 is required for sperm head formation and male fertility.

Purpose: TRANSLIN (TSN) and its binding partner TSNAX have been reported to contribute to a wide spectrum of biological activities including spermatogenesis. TSN accompanies specific mRNA transport in male germ cells through intercellular bridges. A testis-expressed protein TSNAXIP1 was reported to interact with TSNAX. However the role of TSNAXIP1 in spermatogenesis remained unclear. This study aimed to elucidate the role of TSNAXIP1 in spermatogenesis and male fertility in mice. Methods: TSNAXIP1 knockout (KO) mice were generated using the CRISPR-Cas9 system. The fertility, spermatogenesis, and sperm of TSNAXIP1 KO males were analyzed. Results: TSNAXIP1, and especially its domains, are highly conserved between mouse and human. Tsnaxip1 was expressed in testis, but not in ovary. TSNAXIP1 KO mice were generated, and TSNAXIP1 KO males were found to be sub-fertile with smaller testis and lower sperm count. Although no overt abnormalities were observed during spermatogenesis, lack of TSNAXIP1 induced sperm head malformation, resulting in a unique flower-shaped sperm head. Moreover, abnormal anchorage of the sperm neck was frequently observed in TSNAXIP1 null sperm. Conclusion: A testis-expressed gene TSNAXIP1 has important roles in sperm head morphogenesis and male fertility. Moreover, TSNAXIP1 could be a causative gene for human infertility.

Effects of Calyculin a on the Motility and Protein Phosphorylation in Frozen-Thawed Bull Spermatozoa.

In this study, we examined the effects of calyculin A, a phosphatase inhibitor, on motility, protein phosphorylation, and the distribution of phospho-(Ser/Thr) PKA substrates in frozen-thawed bull spermatozoa that are actually used by most farmers for breeding. The data showed that calyculin A, which has been reported to have a positive effect on the motility of ejaculated fresh spermatozoa, distinctly decreased the motility of frozen-thawed bull spermatozoa even if a cell activator, such as caffeine, was present in the incubation medium and that the suppressive effect of calyculin A was dose-dependent and continued for at least 200 min. Immunoblot analyses revealed that de novo protein phosphorylation was not detected in spermatozoa exposed to caffeine or dbcAMP (a cell-permeable cAMP analog), while the addition of calyculin A to the medium brought about the appearance of several phosphorylated proteins at 50 kDa and 75 kDa, suggesting that 50 kDa and 75 kDa proteins, which were phosphorylated by activation of cAMP-dependent PKA, were not dephosphorylated and were accumulated in spermatozoa due to the suppression of calyculin A-sensitive protein phosphatases. Immunofluorescence microscopy revealed that calyculin A caused, alone or in conjunction with caffeine or dbcAMP, the accumulation of phospho-PKA substrates at the annulus, although caffeine or dbcAMP alone did not. This study suggested that calyculin A decreases the motility of frozen-thawed bull spermatozoa concomitant with the accumulation of phospho-(Ser/Thr) PKA substrates at the annulus of flagella.

Novel localizations and interactions of intercellular bridge proteins revealed by proteomic profiling†.

Intercellular bridges (ICBs) connecting germ cells are essential for spermatogenesis, and their deletion causes male infertility. However, the functions and component factors of ICBs are still unknown. We previously identified novel ICB-associated proteins by proteomics analysis using ICB enrichment. Here, we performed immunoprecipitation-proteomics analyses using antibodies specific to known ICB proteins MKLP1, RBM44, and ectoplasmic specialization-associated protein KIAA1210 and predicted protein complexes in the ICB cores. KIAA1210, its binding protein topoisomerase2B (TOP2B), and tight junction protein ZO1 were identified as novel ICB proteins. On the other hand, as well as KIAA1210 and TOP2B, MKLP1 and RBM44, but not TEX14, were localized at the XY body of spermatocytes, suggesting that there is a relationship between ICB proteins and meiotic chromosomes. Moreover, small RNAs interacted with an ICB protein complex that included KIAA1210, RBM44, and MKLP1. These results indicate dynamic movements of ICB proteins and suggest that ICB proteins could be involved not only in the communication between germ cells but also in their epigenetic regulation. Our results provide a novel perspective on the function of ICBs and could be helpful in revealing the biological function of the ICB.

Identification of TMCO2 as an acrosome-associated protein during rat spermiogenesis.

We isolated the transmembrane and coiled-coil domains 2 (Tmco2) gene using a polymerase chain reaction-based subtraction technique. Tmco2 is predominantly expressed in rat testes starting from 4 weeks of age. Rat TMCO2 consists of 187 amino acids with a predicted molecular mass of 20.6 kDa. When expressed in COS7 cells, TMCO2 was found as vesicle-like structures in the cytoplasm, whereas TMCO2ΔTM lacking the transmembrane (TM) region was found diffused in the cytoplasm. These results suggest that the TM region in TMCO2 is essential for its specificity of localization. Immunocytochemical analyzes indicated that rat TMCO2 was localized as small semiluminate bodies or cap-like structures in the vicinity of round spermatid nuclei and as curved lines associated with nuclei of elongated spermatids and caput epididymal spermatozoa. However, it was detected in only a small part of cauda epididymal spermatozoa. Double immunolabeling of the spermatids and spermatozoa with the anti-TMCO2 antibody and the monoclonal anti-MN7 antibody showed that TMCO2 was predominantly associated with the inner acrosomal membrane in spermatids and caput epididymal spermatozoa. Our findings suggest that TMCO2 might be involved in the process of acrosome biogenesis, especially binding of acrosome to a nucleus, during spermiogenesis.

A membrane protein, TMCO5A, has a close relationship with manchette microtubules in rat spermatids during spermiogenesis.

We isolated the transmembrane and coiled-coil domains 5A (Tmco5A) gene using polymerase chain reaction-based subtraction technique and showed that Tmco5A was predominantly expressed in rat testes starting at 4 weeks of postnatal development. When expressed in COS7 cells, TMCO5A was found to be distributed in the endoplasmic reticulum-nuclear membrane (ER-NM) of cells as a membrane-associated protein, while TMCO5AΔC lacking the transmembrane region (TM) mislocalized and diffused throughout the cytoplasm. The result suggested that TM is responsible for the retention of TMCO5A at the ER-NM. Immunocytochemical and immunoblotting analyses indicated that TMCO5A was localized along the posterior part of the nuclei in both round and elongated rat spermatids but disappeared from epididymal spermatozoa. Double immunolabeling of isolated spermatids with the anti-TMCO5A and the anti-ß tubulin antibodies showed that TMCO5A was always found to be closely associated with developing manchette microtubules but did not completely colocalize with them. On the other hand, we found that almost all TMCO5A colocalized with SUN4, a linker of nucleoskeleton and cytoskeleton complex protein present at the posterior part of spermatid nuclei. These data suggested that TMCO5A is located closer to the nuclei than the manchette microtubules. It is likely that TMCO5A, in association with manchette microtubules, is involved in the process of spermiogenesis.

MRG15 is required for pre-mRNA splicing and spermatogenesis.

Splicing can be epigenetically regulated and involved in cellular differentiation in somatic cells, but the interplay of epigenetic factors and the splicing machinery during spermatogenesis remains unclear. To study these interactions in vivo, we generated a germline deletion of MORF-related gene on chromosome 15 (MRG15), a multifunctional chromatin organizer that binds to methylated histone H3 lysine 36 (H3K36) in introns of transcriptionally active genes and has been implicated in regulation of histone acetylation, homology-directed DNA repair, and alternative splicing in somatic cells. Conditional KO (cKO) males lacking MRG15 in the germline are sterile secondary to spermatogenic arrest at the round spermatid stage. There were no significant alterations in meiotic division and histone acetylation. Specific mRNA sequences disappeared from 66 germ cell-expressed genes in the absence of MRG15, and specific intronic sequences were retained in mRNAs of 4 genes in the MRG15 cKO testes. In particular, introns were retained in mRNAs encoding the transition proteins that replace histones during sperm chromatin condensation. In round spermatids, MRG15 colocalizes with splicing factors PTBP1 and PTBP2 at H3K36me3 sites between the exons and single intron of transition nuclear protein 2 (Tnp2). Thus, our results reveal that MRG15 is essential for pre-mRNA splicing during spermatogenesis and that epigenetic regulation of pre-mRNA splicing by histone modification could be useful to understand not only spermatogenesis but also, epigenetic disorders underlying male infertile patients.

Identification of KIAA1210 as a novel X-chromosome-linked protein that localizes to the acrosome and associates with the ectoplasmic specialization in testes.

Cell junctions are necessary for spermatogenesis, and there are numerous types of junctions in testis, such as blood­testis barrier, intercellular bridge, and ectoplasmic specialization (ES). The details of their functions and construction are still unknown. To identify a novel protein essential to the function of a cell junction, we enriched testis membrane protein and analyzed it using a proteomics approach. Here, we report a novel ES protein, which is encoded on the X chromosome and an ortholog of hypothetical human protein KIAA1210. KIAA1210 is expressed in testis predominantly, localized to the sex body in spermatocyte, acrosome, and near ES. Moreover, KIAA1210 possesses a topoisomerase 2 (TOP2)-associated protein PAT1 domain, a herpes simplex virus 1 (HSV-1) large tegument protein UL36 hypothetical domain, and a provisional DNA translocase FtsK domain. Using IP-proteomics with specific antibody to KIAA1210, we identified proteins including TOP2 isoforms as components of a complex with KIAA1210, in cell junctions in testis. The interaction between KIAA1210 and TOP2 was confirmed by two different proteomic analyses. Furthermore, immunofluorescence showed that KIAA1210 and TOP2B co-localize around the sex body in spermatocyte, apical ES, and residual bodies in elongated spermatids. Our findings suggest that KIAA1210 may be essential cell junction protein that interacts with TOP2B to regulate the dynamic change of chromatin structures during spermiogenesis.

Role of satellite cells versus myofibers in muscle hypertrophy induced by inhibition of the myostatin/activin signaling pathway.

Myostatin and activin A are structurally related secreted proteins that act to limit skeletal muscle growth. The cellular targets for myostatin and activin A in muscle and the role of satellite cells in mediating muscle hypertrophy induced by inhibition of this signaling pathway have not been fully elucidated. Here we show that myostatin/activin A inhibition can cause muscle hypertrophy in mice lacking either syndecan4 or Pax7, both of which are important for satellite cell function and development. Moreover, we show that muscle hypertrophy after pharmacological blockade of this pathway occurs without significant satellite cell proliferation and fusion to myofibers and without an increase in the number of myonuclei per myofiber. Finally, we show that genetic ablation of Acvr2b, which encodes a high-affinity receptor for myostatin and activin A specifically in myofibers is sufficient to induce muscle hypertrophy. All of these findings are consistent with satellite cells playing little or no role in myostatin/activin A signaling in vivo and render support that inhibition of this signaling pathway can be an effective therapeutic approach for increasing muscle growth even in disease settings characterized by satellite cell dysfunction.

Regulation of spermatogonial stem cell compartment in the mouse testis.

Spermatogenesis occurs throughout the adult lifetime of males and is supported by a robust stem cell system. Spermatogonial stem cells (SSCs) are the stem cells of postnatal male germ cells, and not only self-renew but also produce differentiated progeny continuously. Recent report revealed that differentiating spermatogonia could revert into an undifferentiated state, although it was believed that SSCs were homogeneous and that differentiating spermatogonia was not reversible. Although several molecules, which regulate SSC, have been identified so far, molecular mechanisms underlying the maintenance of SSCs as well as the reversible developmental lineage of SSCs remain to be elucidated. In this review, we describe a brief overview of spermatogenesis and summarize the molecular regulation of SSC compartment.

Verification of the efficacy of deciduae for determining the genetic type of mouse embryos.

Mouse embryos in the early-implantation stage require manipulation under a microscope. While the extraction of DNA, RNA and proteins from a single sample allows for both determination of genetic type and analysis of gene expression, whole mount analysis is not possible. In this study, we explored the applicability of PCR using extraembryonic tissues, especially the decidual side tissue after isolating the embryos from implantation sites to establish a method for determining the genetic type of embryos. The implantation site was resected at each day from the date of vaginal plug confirmation, separated into embryos and deciduae. Genomic DNA were isolated separately from the embryos and the deciduae. PCR was performed using these genomic DNA, and the band patterns were compared after electrophoresis. As a result, we demonstrated that detecting embryo-derived cells in the decidua allows determination of the sex and presence of transgenes without harming the mouse embryos themselves, from 8.5 days of age. This method enables the determination of the genetic type of mouse embryos without damaging. This technique would expand the adaptations for analysis of mouse implanted embryos.

Abnormal spermatogenesis and reduced fertility in transgenic mice expressing the immediate-early protein IE180 of pseudorabies virus.

Transcription factors of alphaherpesviruses not only control the expression of their own viral genes, but also influence the gene expression of mammalian cells. In the course of breeding of the transgenic mouse line (TgIE96) expressing the immediate-early protein IE180 of pseudorabies virus belonging to the subfamily Alphaherpesvirinae, we found that TgIE96 male mice suffered from severe breeding difficulties. Testes of TgIE96 were smaller than that of non-transgenic littermates and abnormal spermatogenesis such as morphological, numerical and functional anomalies of spermatozoa were found in the transgenic mouse line. Expression of IE180 was detected in the germ cells at all stages, especially spermatocytes, and fewer Sertoli cells. In addition, expression of IE180 was also detected in the germinal cells of C57BL/6 mice inoculated with PRV into their testes. These results suggest that IE180 of PRV induces male infertility by abnormal spermatogenesis, which effect morphological, numerical, and functional anomalies of spermatozoa, in transgenic mice.

ProSE-Pero: Peroxisomal Protein Localization Identification Model Based on Self-Supervised Multi-Task Language Pre-Training Model.

BACKGROUND: Peroxisomes are membrane-bound organelles that contain one or more types of oxidative enzymes. Aberrant localization of peroxisomal proteins can contribute to the development of various diseases. To more accurately identify and locate peroxisomal proteins, we developed the ProSE-Pero model. METHODS: We employed three methods based on deep representation learning models to extract the characteristics of peroxisomal proteins and compared their performance. Furthermore, we used the SVMSMOTE balanced dataset, SHAP interpretation model, variance analysis (ANOVA), and light gradient boosting machine (LightGBM) to select and compare the extracted features. We also constructed several traditional machine learning methods and four deep learning models to train and test our model on a dataset of 160 peroxisomal proteins using tenfold cross-validation. RESULTS: Our proposed ProSE-Pero model achieves high performance with a specificity (Sp) of 93.37%, a sensitivity (Sn) of 82.41%, an accuracy (Acc) of 95.77%, a Matthews correlation coefficient (MCC) of 0.8241, an F1 score of 0.8996, and an area under the curve (AUC) of 0.9818. Additionally, we extended our method to identify plant vacuole proteins and achieved an accuracy of 91.90% on the independent test set, which is approximately 5% higher than the latest iPVP-DRLF model. CONCLUSIONS: Our model surpasses the existing In-Pero model in terms of peroxisomal protein localization and identification. Additionally, our study showcases the proficient performance of the pre-trained multitasking language model ProSE in extracting features from protein sequences. With its established validity and broad generalization, our model holds considerable potential for expanding its application to the localization and identification of proteins in other organelles, such as mitochondria and Golgi proteins, in future investigations.

Disease-related compound identification based on deeping learning method.

Acute lung injury (ALI) is a serious respiratory disease, which can lead to acute respiratory failure or death. It is closely related to the pathogenesis of New Coronavirus pneumonia (COVID-19). Many researches showed that traditional Chinese medicine (TCM) had a good effect on its intervention, and network pharmacology could play a very important role. In order to construct "disease-gene-target-drug" interaction network more accurately, deep learning algorithm is utilized in this paper. Two ALI-related target genes (REAL and SATA3) are considered, and the active and inactive compounds of the two corresponding target genes are collected as training data, respectively. Molecular descriptors and molecular fingerprints are utilized to characterize each compound. Forest graph embedded deep feed forward network (forgeNet) is proposed to train. The experimental results show that forgeNet performs better than support vector machines (SVM), random forest (RF), logical regression (LR), Naive Bayes (NB), XGBoost, LightGBM and gcForest. forgeNet could identify 19 compounds in Erhuang decoction (EhD) and Dexamethasone (DXMS) more accurately.

The testis-enriched histone demethylase, KDM4D, regulates methylation of histone H3 lysine 9 during spermatogenesis in the mouse but is dispensable for fertility.

Epigenetic modifications, and methylation of histones in particular, dynamically change during spermatogenesis. Among various methylations of histone H3, methylation of histone H3 lysine 9 (H3K9) and its regulation are essential for spermatogenesis. Trimethytransferases as well as dimethyltransferase are required for meiotic progression. In addition, didemethylase of H3K9 is also critical for spermatogenesis through transcriptional regulation of spermatid-specific genes. However, the requirement for demethylation of trimethylated H3K9 (H3K9me3) during spermatogenesis remains to be elucidated. Here, we report the targeted disruption of KDM4D, a testis-enriched tridemethylase of H3K9. Kdm4d-null mice are viable and fertile and do not show any obvious phenotype. However, H3K9me3 accumulates significantly in Kdm4d-null round spermatids, and the distribution of methylated H3K9 in germ cells is dramatically changed. Nevertheless, the progression of spermatogenesis and the number of spermatozoa are normal, likely secondary to the earlier nuclear localization of another H3K9 tridemethylase, KDM4B, in Kdm4d-null elongating spermatids. These results suggest that demethylation of H3K9me3 in round spermatids is dispensable for spermatogenesis but that possible defects in Kdm4d-null elongating spermatids could be rescued by functional redundancy of the KDM4B demethylase.

Comparative distributions of RSBN1 and methylated histone H4 Lysine 20 in the mouse spermatogenesis.

During spermatogenesis, nuclear architecture of male germ cells is dynamically changed and epigenetic modifications, in particular methylation of histones, highly contribute to its regulation as well as differentiation of male germ cells. Although several methyltransferases and demethylases for histone H3 are involved in the regulation of spermatogenesis, roles of either histone H4 lysine 20 (H4K20) methyltransferases or H4K20 demethylases during spermatogenesis still remain to be elucidated. Recently, RSBN1 which is a testis-specific gene expressed in round spermatids was identified as a demethylase for dimethyl H4K20. In this study, therefore, we confirm the demethylase function of RSBN1 and compare distributions between RSBN1 and methylated H4K20 in the seminiferous tubules. Unlike previous report, expression analyses for RSBN1 reveal that RSBN1 is not a testis-specific gene and is expressed not only in round spermatids but also in elongated spermatids. In addition, RSBN1 can demethylate not only dimethyl H4K20 but also trimethyl H4K20 and could convert both dimethyl H4K20 and trimethyl H4K20 into monomethyl H4K20. When distribution pattern of RSBN1 in the seminiferous tubule is compared to that of methylated H4K20, both dimethyl H4K20 and trimethyl H4K20 but not monomethyl H4K20 are disappeared from RSBN1 positive germ cells, suggesting that testis-specific distribution patterns of methylated H4K20 might be constructed by RSBN1. Thus, novel expression and function of RSBN1 could be useful to comprehend epigenetic regulation during spermatogenesis.

Fasting-induced FGF21 signaling activates hepatic autophagy and lipid degradation via JMJD3 histone demethylase.

Autophagy is essential for cellular survival and energy homeostasis under nutrient deprivation. Despite the emerging importance of nuclear events in autophagy regulation, epigenetic control of autophagy gene transcription remains unclear. Here, we report fasting-induced Fibroblast Growth Factor-21 (FGF21) signaling activates hepatic autophagy and lipid degradation via Jumonji-D3 (JMJD3/KDM6B) histone demethylase. Upon FGF21 signaling, JMJD3 epigenetically upregulates global autophagy-network genes, including Tfeb, Atg7, Atgl, and Fgf21, through demethylation of histone H3K27-me3, resulting in autophagy-mediated lipid degradation. Mechanistically, phosphorylation of JMJD3 at Thr-1044 by FGF21 signal-activated PKA increases its nuclear localization and interaction with the nuclear receptor PPARα to transcriptionally activate autophagy. Administration of FGF21 in obese mice improves defective autophagy and hepatosteatosis in a JMJD3-dependent manner. Remarkably, in non-alcoholic fatty liver disease patients, hepatic expression of JMJD3, ATG7, LC3, and ULK1 is substantially decreased. These findings demonstrate that FGF21-JMJD3 signaling epigenetically links nutrient deprivation with hepatic autophagy and lipid degradation in mammals.

Putative "stemness" gene jam-B is not required for maintenance of stem cell state in embryonic, neural, or hematopoietic stem cells.

Many genes have been identified that are specifically expressed in multiple types of stem cells in their undifferentiated state. It is generally assumed that at least some of these putative "stemness" genes are involved in maintaining properties that are common to all stem cells. We compared gene expression profiles between undifferentiated and differentiated embryonic stem cells (ESCs) using DNA microarrays. We identified several genes with much greater signal in undifferentiated ESCs than in their differentiated derivatives, among them the putative stemness gene encoding junctional adhesion molecule B (Jam-B gene). However, in spite of the specific expression in undifferentiated ESCs, Jam-B mutant ESCs had normal morphology and pluripotency. Furthermore, Jam-B homozygous mutant mice are fertile and have no overt developmental defects. Moreover, we found that neural and hematopoietic stem cells recovered from Jam-B mutant mice are not impaired in their ability to self-renew and differentiate. These results demonstrate that Jam-B is dispensable for normal mouse development and stem cell identity in embryonic, neural, and hematopoietic stem cells.