Search Results (469)

The wnt gene family encodes a group of highly conserved secreted glycoproteins that play essential roles in vertebrate development, including tissue patterning, cell differentiation, and gonadal regulation. However, the genomic organization, evolutionary dynamics, and functional roles of Wnt signaling components in flatfish remain poorly understood. In this study, we performed a comprehensive genome-wide identification, evolutionary characterization, expression profiling, and functional analysis of wnt genes in Cynoglossus semilaevis, a flatfish species exhibiting ZW/ZZ sex determination and temperature-induced sex reversal. A total of 20 wnt genes were identified and classified into 13 subfamilies, displaying conserved structural organization and phylogenetic relationships consistent with other teleosts. Chromosomal mapping revealed lineage-specific WNT clusters, including a unique wnt3–wnt7b–wnt5b–wnt16 block, as well as syntenic associations with reproduction-related genes (e.g., adipor2, sema3a, nape-pld, erc2, lamb2), suggesting coordinated genomic regulation. Tissue transcriptome analysis demonstrated strong sex- and tissue-biased expression patterns, with wnt5a predominantly expressed in ovaries and wnt5b specifically upregulated in pseudo-male testes. Functional assays revealed that knockdown of wnt5a or wnt5b induced testis-specific genes (sox9b, tesk1) and suppressed ovarian markers (foxl2, cyp19a1a), indicating antagonistic regulatory roles in gonadal fate determination. Promoter analysis identified yy1a as a selective repressor of wnt5b, but not wnt5a, providing a mechanistic basis for paralog divergence. Furthermore, pull-down combined with LC–MS/MS analysis showed that WNT5b interacts with proteins enriched in ribosome biogenesis and ubiquitin-mediated proteolysis, suggesting a role in translational regulation and protein turnover during spermatogenesis. Together, these findings establish WNT5 signaling—particularly wnt5b—as a key driver of testicular development in C. semilaevis and provide new insights into the molecular mechanisms underlying sex differentiation and sex reversal in flatfish. Full article

Glutamine synthetase, a critical enzyme catalyzing the conversion of glutamate and ammonia into glutamine, has been shown to influence sperm development in mammals. Here, we carried out functional analysis of Bombyx mori homolog of glutamine synthetase 1 (BmGS1) and screened its small-molecule inhibitor. RT-PCR and qPCR showed that BmGS1 was specifically expressed in the testis of the silkworm, with the highest expression in the moth stage. Subcellular localization revealed that the BmGS1 protein was localized in mitochondria and cytoplasm. Identification of upstream regulatory factors revealed that the expression of BmGS1 is positively regulated by the sex-related transcription factor Bmdsx. Virtual screening, molecular docking and MD simulations showed that the small molecule Ethylhexyl triazone (ET), as well as the known GS inhibitor L-Methionine -DL-sulfoximine (MSX), could be stably bound to BmGS1. Subsequently, site-specific mutation and fluorescence binding assays revealed that the putative key sites of ET binding to the protein were E79 and R265, and the putative key sites of MSX binding to the protein were E81, R245, and R286. Both in vitro and in vivo experiments demonstrated that inhibitor treatment significantly attenuated BmGS1 enzymatic activity. Inhibitor-injected silkworms showed reduced fertilization rates compared to control groups. Our findings raise BmGS1 as a potential target for silkworm sterility. Full article

Allotetraploid fish produced by distant hybridization are valuable germplasm for the mass production of sterile triploids. The allotetraploid crucian–carp hybrid (4nAT, 4n = 200) is derived from the intergeneric cross between a female red crucian carp (Carassius auratus red var., 2n = 100) and a male common carp (Cyprinus carpio L., 2n = 100). However, after 33 successive generations, this lineage faces a critical bottleneck in maintaining male fertility. The present study aimed to develop new biomarkers for testicular development and characterize the associated functional gene expression profile in 4nAT. Following whole-genome resequencing and selection signature analysis of 15 male 4nAT individuals from each of the high-development group (HDG) and low-development group (LDG), ZSWIM7 (Zinc Finger SWIM-Type Containing 7), a gene implicated in reproductive development, was selected as a candidate for further fertility association studies. Seven SNPs were screened in the coding region of ZSWIM7 of 70 4nAT males; among these, SNP3 (c.23T/C) exhibited a significant correlation between genotypes and testicular development: individuals with the CT genotype showed a higher gonadosomatic index (1.17 ± 0.68 vs. 0.65 ± 0.50) and greater counts of mature spermatozoa (2537.67 ± 283.95 vs. 341.56 ± 121.66) compared to those with the TT genotype. Further quantitative PCR and immunofluorescence assays demonstrated that ZSWIM7 was highly expressed in the testis and specifically localized to the nuclei of early meiotic primary spermatocytes. Collectively, these results establish ZSWIM7 as a promising biomarker for 4nAT testicular development, offering a potential molecular tool for maintaining male fertility in allotetraploid fish breeding. Full article

The World Health Organization estimates that about 15% of couples in their adult years in industrialized countries experience infertility, which is described as the inability of a sexually active and non-contraceptive couple to achieve spontaneous pregnancy within a year. Environmental pollution is a significant health concern worldwide and one of the possible risk factors leading to male infertility. Cadmium is a common heavy toxin derived from industrial activities, a ubiquitous environmental pollutant, and can cause severe harm to various organs including the testis. Cadmium toxicity can lead to severe impairment of male germ cells in both rodents and humans, which can result in azoospermia. The negative effects of cadmium on the testicles are caused by its induction of oxidative stress, spermatogenic apoptosis, and testicular inflammation or its detriment to androgenic and sperm cell functions, which damages the vascular endothelium and blood–testis barrier. Overall, this review describes the detrimental impact of cadmium on the testicles and its effect on male infertility. Therefore, by considering recent research findings and identifying future research directions, this review underlines the need to develop new treatments for male infertility related to heavy metal exposure. Full article

Thamnaconus septentrionalis is an emerging commercially important aquaculture species in China, distributed extensively in the Indo-West Pacific Ocean. Recently, because of the seriously declining population and considerable economic potential of this aquaculture fish, increasing attention has been paid to the conservation and development of T. septentrionalis. Artificial fish breeding is essential and has become progressively implemented on local farms in China, which benefits the protection of T. septentrionalis resources and facilitates the development of its fishery industry. Previous studies have demonstrated that temperature could significantly influence ovary development in T. septentrionalis. However, the potential molecular mechanisms underlying the influence of temperature on testis development in T. septentrionalis have been scarcely studied. Thus, this study comprehensively explores the effects of temperature on testis development in T. septentrionalis using histological observation and transcriptomic techniques. Histological and transmission electron microscopy analyses indicated that T. septentrionalis testes, undergoing a rise in temperature, developed from phase III to IV. Transcriptomic analysis identified 315 differentially expressed genes, including 200 upregulated and 115 downregulated genes. Moreover, rising temperatures may enhance testis development by regulating steroid hormone biosynthesis, cellular senescence, and nucleotide metabolism. The upregulation of four genes (hsd11b2, cyp11b, cyp11a, and hsd17b3) involved in the steroid hormone biosynthesis process may significantly contribute to the increased level of testosterone and 11-keto-testosterone. This study is the first to elucidate the potential molecular mechanism involved in T. septentrionalis testis development induced by temperature, offering valuable and novel insights for its artificial breeding and fishery resources conservation. Full article

Ovarian cancer (OC) remains the leading cause of death among gynecologic cancers. Most women diagnosed with OC at advanced stages eventually develop relapse and chemoresistance, leading to poor clinical outcomes. While piRNAs have emerged as critical regulators of gene expression and tumor biology, their specific roles in OC remain to be fully elucidated. This study integrated clinical and computational analyses to investigate the expression pattern and functional relevance of P-element-induced wimpy testis (PIWI)-interacting RNA-823 (piR-823) and its associated protein piwi-like RNA-mediated gene silencing 1 (PIWIL1)/DNA methyltransferase 3B (DNMT3B)/E-cadherin (CDH1) axis in OC tissues from 40 patients, with 20 non-cancer control samples. Expression profiling was performed using qPCR on OC and normal ovarian tissues, followed by correlation and regression analyses. Public databases, including GEPIA, TNM plot, and MethBank, were explored to validate gene expression, methylation status, and pathway enrichment. Our results revealed that piR-823, PIWIL1, and DNMT3B were significantly upregulated in OC tissues (p < 0.001, p = 0.009, and p < 0.001, respectively), and they correlated positively with each other and inversely with CDH1 expression. CDH2, OCT4, and NANOG were significantly upregulated (p = 0.011, p = 0.03, and p < 0.001, respectively), whereas CDH1 expression was significantly downregulated (p < 0.001) in OC tissues. In silico analyses supported DNMT3B-mediated CDH1 promoter methylation, epithelial–mesenchymal transition (EMT), and stemness pathway enrichment. Our integrated computational and clinical analyses indicate that the piR-823/PIWIL1/DNMT3B/CDH1 axis is a putative epigenetic regulator of EMT and cancer stemness in ovarian cancer. Additionally, piR-823 may serve as a promising prognostic biomarker and therapeutic target, offering novel insights into OC pathogenesis and treatment. Full article

Hemibagrus guttatus is a large omnivorous fish of significant economic value, listed as a Class II protected species in the National Key Protected Wildlife List in 2021 in China. To provide a theoretical foundation for the artificial breeding of H. guttatus, this study employs high-throughput transcriptome sequencing of testes and ovaries to elucidate the molecular regulatory pathways involved in sex differentiation. Because H. guttatus exhibits no obvious sexual dimorphism even during the breeding season, the distinctive contribution of this study compared with previous gonadal-transcriptomic investigations in other Siluriformes lies not only in documenting sex-biased genes but also in providing a molecular foundation for developing non-lethal sex-identification methods for this morphologically indistinguishable species. A total of 303,192,896 raw reads were obtained, with an effective data rate of 98.4%, indicating high sequencing quality. Differential expression analysis identified 8694 genes, including 6369 upregulated in testes and 2325 upregulated in ovaries. Among these, 88 genes were functionally annotated as sex-related, with 62 testis-biased genes such as spata17, sox9, and dmrt1, and 26 ovary-biased genes including cyp19a, wnt8, and sox12. KEGG pathway enrichment analysis revealed that the TGF-β signaling pathway, insulin secretion, and steroid hormone biosynthesis may play crucial roles in gonadal development and differentiation in H. guttatus. The expression patterns of key genes such as hsd11b1, amh, and insl3 were validated by quantitative real-time PCR, showing consistency with the transcriptome results. These findings lay a molecular foundation for understanding the regulatory mechanisms of sex differentiation in H. guttatus, and provide candidate genes for further investigation into the genetic basis of gonadal development, which is essential for improving artificial reproduction and selective breeding practices. Full article

Transforming growth factor β (TGF-β) superfamily members are critical in teleost sex determination and differentiation. Tgfb2b is an important TGF-β ligand gene exhibiting dominant expression in the ovary of Chinese tongue sole (Cynoglossus semilaevis), yet its function in sex regulation remains unclear. In the present study, the gene expression pattern, transcriptional regulation, and knockdown effect were examined. Its expression persisted and showed a gradual increase throughout ovarian development from 3 months to 1.5 years post-hatching. In situ hybridization (ISH) revealed that the gene was distributed across oocytes at stages I–III, while scarcely detectable in the testis. The transcriptional factors CCAAT/enhancer binding protein α (C/EBPα) and Jun proto-oncogene AP-1 transcription factor subunit (c-Jun) could repress the activity of tgfb2b promoter. In vitro knockdown of tgfb2b in C. semilaevis ovarian cells led to downregulation of its downstream genes (e.g., smad1 and smad2) as well as other sex-related genes (e.g., foxl2 and esr2b). Moreover, multi-omics analysis indicated that, in C. semilaevis gonads, a miRNA named novel-m0083-3p showed an opposite expression pattern with tgfb2b and might have a binding site with the gene. By dual-luciferase assay, tgfb2b was validated to be directly targeted and suppressed by the miRNA. These results demonstrate that tgfb2b plays a significant role in ovarian differentiation and development. Further functional and molecular studies on the interplay between tgfb2b and the foxl2–cyp19a–esr axis will help elucidate the regulatory network underlying sex development in teleost. Full article

Background: Testicular dysgenesis syndrome (TDS) is a complex disorder of the male reproductive system related to disfunction of the fetal testis. The clinical features of TDS may be evident at birth or infancy (cryptorchidism, hypospadias and/or reduced anogenital distance) or occur later in adulthood (testis cancer, infertility). Genetic background seems to be important for genetic predisposition, with new genes being associated with components of the syndrome in last years. Interestingly, the incidence of clinical manifestations of TDS has been increasing in many countries in recent decades, suggesting that genetic predisposition alone cannot explain this trend. Consequently, the hypothesis of multifactorial etiopathogenesis is becoming increasingly accepted nowadays, with environmental factors probably acting during early developmental stages in genetically predisposed individuals. Methods: In this narrative review, we aim to critically evaluate genetic and non-genetic factors involved in the pathogenesis of TDs. Results: Important associations with intrauterine growth disorders and maternal diseases (overweight/obesity and diabetes) as well as lifestyle factors (e.g., smoking and alcohol abuse) were found. In such context, endocrine disruptors probably play a major role. These substances are widely used in industry and can exert estrogenic and antiandrogenic effects, potentially interfering with the development of the fetal gonad. Conclusions: Considering their possible impact on male sexual health, more attention should be focused on maternal modifiable factors to confirm with prospective studies the mixed results of available evidence. Full article

Arginine vasotocin (AVT) has recently emerged as a local regulator of testicular function in fish. Using ex vivo culture system, it was demonstrated that AVT directly stimulates androgen-dependent basal spermatogenesis in zebrafish. In the presence of gonadotropins, AVT enhanced FSH-induced development of early phases of spermatogonial proliferation while blocking FSH-mediated spermiogenesis. However, AVT promoted proliferation of LH-induced pre-meiotic and meiotic germ cell populations without affecting the final stages of spermiogenesis. These findings led to the hypothesis that AVT plays a role by promoting early germ cell proliferation and differentiation while simultaneously inhibiting premature progression through spermiogenesis. To test this hypothesis, we investigated the chronic effects of AVT on adult zebrafish testes, in vivo. Prolonged AVT treatment for 21 days led to dose-dependent accumulation of undifferentiated type A spermatogonia and reduced post-meiotic germ cells and spermatozoa. We also observed decreased plasma 11-ketotestosterone (11-KT) levels and downregulation of fshr. This was accompanied by a basal suppression of avt and its receptors, avpr1aa, avpr1ab, avpr2aa, avpr2ab, avpr2l, in both brain and testis during the pre-spawning phase. The present findings, along with those of previously published studies, collectively demonstrate that AVT presence during the early stages of testicular development promotes spermatogonia proliferation while diminishing FSH-induced premature progress toward spermatogenesis. This occurs until later stages, when AVT expression is diminished, allowing for optimal LH-induced spermiogenesis in zebrafish. Full article

Background: Hypospadias development is influenced by prenatal androgen levels, with genetic factors typically playing a significant role. Through whole-exome sequencing, we found that rare damaging variants in DNAH8 (dynein axonemal heavy chain 8) were significantly enriched in hypospadias cases. However, the role of DNAH8 deficiency in hypospadias pathogenesis remains unclear. Objectives: This study aimed to clarify the function of DNAH8 in urethral development and fusion. Materials and Methods: Using CRISPR/Cas9, we generated DNAH8 knockout mice and employed a multi-disciplinary approach to evaluate urogenital development, male differentiation, testosterone levels, steroid biosynthesis gene expression, and cellular changes in fetal testes and external genitalia. Results:DNAH8 knockout mice presented abnormal masculinization phenotype, and fetal mice exhibited urethral fusion defects and hypoplastic glans during early urethral development. DNAH8 knockout was found to reduce prenatal testosterone levels and steroid biosynthesis in the testes. Based on single-cell sequencing and multicolor immunofluorescence, we demonstrated that in the early stage of fetal testis development, the loss of DNAH8 function affected the differentiation of Sertoli and steroidogenic cell lineages, thereby impairing testosterone synthesis ability during the masculinization programming window. Meanwhile, we identified two key distal glans cell populations that cause abnormal urethral fusion and hypoplastic glans. Furthermore, DNAH8 knockout could synergistically interact with low-dose endocrine-disrupting chemicals, increasing the incidence of urethral fusion defects at E16.5, and led to clear hypospadias phenotypes at E18.5. Conclusions: Loss of DNAH8 delays differentiation of Sertoli and steroidogenic lineages, reduces prenatal testosterone, and, with environmental exposure, increases hypospadias risk. Full article

46,XY gonadal dysgenesis, characterised by absent or defective testicular development in individuals with a 46,XY karyotype, results from disruptions in the genetic programme governing testis determination and differentiation during embryogenesis. While monogenic causes explain approximately 50% of cases, emerging evidence suggests an oligogenic basis in some patients. However, many cases remain without a definitive molecular diagnosis. In this study, we investigated a patient with 46,XY gonadal dysgenesis to explore the underlying genetic aetiology. Whole-exome sequencing in the patient did not reveal any pathogenic variants in genes previously associated with this condition. Instead, it detected rare variants in STARD9 and CDK5RAP2, which encode centrosomal proteins known to interact with each other. Gene expression analysis of embryonic gonads revealed that STARD9 is sexually dimorphic, with the highest expression in testis-specific Sertoli cells, while CDK5RAP2 is ubiquitously expressed, including in Sertoli cells. These findings suggest a role for both genes in Sertoli cell development, implicating them in the pathogenesis of 46,XY gonadal dysgenesis. To evaluate the functional relevance of the identified variants, we performed molecular dynamics simulations, which suggest that these variants may impair the individual and/or combined functions of STARD9 and CDK5RAP2 proteins. This study is the first to propose a role for STARD9 and CDK5RAP2 genes in human Sertoli cell development and highlights their potential contribution to 46,XY gonadal dysgenesis. Full article

Gene expression plays a fundamental role in defining the characteristics of living organisms. To deepen our understanding of tissue-specific gene expression, we analyzed transcript variant enrichment across different tissues in human and Drosophila melanogaster. Datasets are widely accessible for both of these organisms. Given the substantial volume of available information, we have focused our interest on three fundamentally distinct tissues: the brain, where both neuronal and glial cells exhibit a relatively high cellular surface area, thus requiring a large amount of lipids; the adipose tissue, which is well-known for lipid storage; and the testis, which contains a massive number of developing spermatids with high membrane requirement. These three organs have fundamental differences in their structure and function yet share some common features; they all have lipid-rich cells and have special metabolic pathways. Most studies focus on gene expression, and transcript level analyses are less common; therefore, we aimed to characterize the transcript profiles of these tissues and examine evolutionarily conserved pathways between humans and Drosophila. Additionally, we analyzed the flanking sequences of transcriptional start sites of tissue-enriched transcripts. Our findings suggest that Drosophila tissues exhibit more distinct regulation of gene expression in individual tissues (weaker correlation in expression and variable nucleotide content in core promoter), whereas human gene expression is more generalized, likely relying more heavily on distal regulatory elements for tissue-specific expression. Through network analysis, summarizing tissue specificity, physical interactions, and orthologue data, we identified shared central pathways among these tissues. A relatively large network was observable in the testis, where the ubiquitin proteasome system, various kinases and transcription factors showed central position in both organisms. Additionally, we highlighted the evolutionary potential of highly enriched testis-specific transcripts. This work provides valuable insights into the mechanisms underlying tissue-specific gene expression and evolutionary conservation. Full article

Human T-lymphotropic virus type 1 (HTLV-1) infection is associated with a spectrum of clinical outcomes, ranging from lifelong asymptomatic carriage to severe conditions such as HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) and adult T-cell leukemia/lymphoma (ATLL). Although antibody responses are known to shape immune regulation, the functional relevance of IgG idiotype repertoires in HTLV-1 pathogenesis remains poorly understood. This study investigated the immunomodulatory effects of IgG from individuals with distinct HTLV-1 clinical outcomes. IgG was purified from pooled serum samples of asymptomatic carriers (ACs), HAM/TSP, and ATLL patients and used to stimulate peripheral blood mononuclear cells (PBMCs) from healthy donors. Cytokine production in CD4⁺, CD8⁺, and γδ T cells was assessed by flow cytometry. Additionally, proteome-wide IgG reactivity was evaluated using a human protein microarray encompassing over 21,000 proteins, and bioinformatic analyses were conducted to identify protein–protein interaction networks and tissue-specific autoreactivity. HAM/TSP-derived IgG selectively enhanced IFN-γ production in all T-cell subsets and suppressed IL-4 in CD4⁺ T cells. ATLL-derived IgG induced IL-9 and IL-13 production in CD4⁺ T cells, and both HAM/TSP and ATLL IgG elevated IL-13 levels in CD8⁺ T cells. Microarray data revealed distinct autoreactive IgG profiles across clinical groups, targeting immune-related proteins, apoptotic regulators, and proteins expressed in T cells, monocytes, and non-immune tissues such as brain and testis. Notably, no functional or structural clustering was observed in protein–protein interaction networks, suggesting these reactivities reflect complex, idiotype-specific immune alterations rather than compensatory responses. The present findings suggest that HTLV-1 infection may be associated with the development of distinct IgG repertoires that potentially modulate cytokine responses and exhibit broad reactivity toward human proteins. Such patterns could contribute to immune dysregulation and may partially explain the divergent clinical trajectories observed in HAM/TSP and ATLL. Further investigations are warranted to validate these observations at the individual level and to clarify their mechanistic relevance in disease progression. Full article

Background/Objectives: Long non-coding RNA (lncRNA) was structurally similar to mRNAs, yet they could not be translated into proteins. While an increasing number of reports have systematically identified and described lncRNA in model species, information about non-model species remains scarce. Sea cucumber Holothuria leucospilota could be used for both medicinal and food purposes, which have high economic value, gradually attracting the attention of researchers. Methods: In this research, we constructed lncRNA library and compared the difference in lncRNA expression profiles between testis and ovary of sea cucumber H. leucospilota. To elucidate the molecular interactions between lncRNA and mRNA, we computationally predicted potential complementary binding sites through analysis of both cis- and trans-acting antisense mechanisms. Subsequent Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses demonstrated that the identified target genes are potentially involved in the regulatory pathways governing gonad development. Results: Quantitative reverse transcription PCR analysis showed that MSTRG.32831.1-sox9 and MSTRG.57315.1-mthfr exhibited a high expression pattern in testis; while MSTRG.11041.1-mafa and MSTRG.11074.1-macf1 showed a high expression pattern in the ovary. Conclusions: Deciphering lncRNA–mRNA expression patterns may uncover fundamental principles governing reproductive regulation in marine invertebrates. This discovery not only deepens understanding in this field but also provides valuable comparative insights for developmental biology. Full article