RESUMO
In mammals, the transition from mitosis to meiosis facilitates the successful production of gametes. However, the regulatory mechanisms that control meiotic initiation remain unclear, particularly in the context of complex histone modifications. Herein, we show that KDM2A, acting as a lysine demethylase targeting H3K36me3 in male germ cells, plays an essential role in modulating meiotic entry and progression. Conditional deletion of Kdm2a in mouse pre-meiotic germ cells results in complete male sterility, with spermatogenesis ultimately arrested at the zygotene stage of meiosis. KDM2A deficiency disrupts H3K36me2/3 deposition in c-KIT+ germ cells, characterized by a reduction in H3K36me2 but a dramatic increase in H3K36me3. Furthermore, KDM2A recruits the transcription factor E2F1 and its co-factor HCFC1 to the promoters of key genes required for meiosis entry and progression, such as Stra8, Meiosin, Spo11, and Sycp1. Collectively, our study unveils an essential role for KDM2A in mediating H3K36me2/3 deposition and controlling the programmed gene expression necessary for the transition from mitosis to meiosis during spermatogenesis.
RESUMO
Spermatogenesis depends on the crosstalk of Sertoli cells (SCs) and germ cells. However, the gene regulatory network establishing the communications between SCs and germ cells remains unclear. Here, we report that heterogeneous nuclear ribonucleoprotein H1 (hnRNPH1) in SCs is essential for the establishment of crosstalk between SCs and germ cells. Conditional knockout of hnRNPH1 in mouse SCs leads to compromised blood-testis barrier function, delayed meiotic progression, increased germ cell apoptosis, sloughing of germ cells and, eventually, infertility of mice. Mechanistically, we discovered that hnRNPH1 could interact with the splicing regulator PTBP1 in SCs to regulate the pre-mRNA alternative splicing of the target genes functionally related to cell adhesion. Interestingly, we also found hnRNPH1 could cooperate with the androgen receptor, one of the SC-specific transcription factors, to modulate the transcription level of a group of genes associated with the cell-cell junction and EGFR pathway by directly binding to the gene promoters. Collectively, our findings reveal a crucial role for hnRNPH1 in SCs during spermatogenesis and uncover a potential molecular regulatory network involving hnRNPH1 in establishing Sertoli-germ cell crosstalk.
Assuntos
Células de Sertoli , Espermatogênese , Animais , Masculino , Camundongos , Fertilidade/fisiologia , Células Germinativas/metabolismo , Ribonucleoproteínas Nucleares Heterogêneas/genética , Ribonucleoproteínas Nucleares Heterogêneas/metabolismo , Camundongos Knockout , Proteína de Ligação a Regiões Ricas em Polipirimidinas/genética , Proteína de Ligação a Regiões Ricas em Polipirimidinas/metabolismo , Células de Sertoli/metabolismo , Espermatogênese/genética , Testículo/metabolismo , Fatores de Transcrição/metabolismoRESUMO
Teratozoospermia is a significant cause of male infertility, but the pathogenic mechanism of acephalic spermatozoa syndrome (ASS), one of the most severe teratozoospermia, remains elusive. We previously reported Spermatogenesis Associated 6 (SPATA6) as the component of the sperm head-tail coupling apparatus (HTCA) required for normal assembly of the sperm head-tail conjunction, but the underlying molecular mechanism has not been explored. Here, we find that the co-chaperone protein BAG5, expressed in step 9-16 spermatids, is essential for sperm HTCA assembly. BAG5-deficient male mice show abnormal assembly of HTCA, leading to ASS and male infertility, phenocopying SPATA6-deficient mice. In vivo and in vitro experiments demonstrate that SPATA6, cargo transport-related myosin proteins (MYO5A and MYL6) and dynein proteins (DYNLT1, DCTN1, and DNAL1) are misfolded upon BAG5 depletion. Mechanistically, we find that BAG5 forms a complex with HSPA8 and promotes the folding of SPATA6 by enhancing HSPA8's affinity for substrate proteins. Collectively, our findings reveal a novel protein-regulated network in sperm formation in which BAG5 governs the assembly of the HTCA by activating the protein-folding function of HSPA8.
Assuntos
Proteínas do Citoesqueleto , Infertilidade Masculina , Teratozoospermia , Tiazóis , Animais , Humanos , Masculino , Camundongos , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Dineínas/metabolismo , Proteínas de Choque Térmico HSC70/genética , Proteínas de Choque Térmico HSC70/metabolismo , Infertilidade Masculina/genética , Infertilidade Masculina/patologia , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Dobramento de Proteína , Sêmen/metabolismo , Cabeça do Espermatozoide/fisiologia , Espermatogênese/genética , Espermatozoides/metabolismo , Teratozoospermia/metabolismo , Teratozoospermia/patologiaRESUMO
In our prior investigation, we discerned loss-of-function variants within the gene encoding glutamine-rich protein 2 (QRICH2) in two consanguineous families, leading to various morphological abnormalities in sperm flagella and male infertility. The Qrich2 knockout (KO) in mice also exhibits multiple morphological abnormalities of the flagella (MMAF) phenotype with a significantly decreased sperm motility. However, how ORICH2 regulates the formation of sperm flagella remains unclear. Abnormal glutamylation levels of tubulin cause dysplastic microtubules and flagella, eventually resulting in the decline of sperm motility and male infertility. In the current study, by further analyzing the Qrich2 KO mouse sperm, we found a reduced glutamylation level and instability of tubulin in Qrich2 KO mouse sperm flagella. In addition, we found that the amino acid metabolism was dysregulated in both testes and sperm, leading to the accumulated glutamine (Gln) and reduced glutamate (Glu) concentrations, and disorderly expressed genes responsible for Gln/Glu metabolism. Interestingly, mice fed with diets devoid of Gln/Glu phenocopied the Qrich2 KO mice. Furthermore, we identified several mitochondrial marker proteins that could not be correctly localized in sperm flagella, which might be responsible for the reduced mitochondrial function contributing to the reduced sperm motility in Qrich2 KO mice. Our study reveals a crucial role of a normal Gln/Glu metabolism in maintaining the structural stability of the microtubules in sperm flagella by regulating the glutamylation levels of the tubulin and identifies Qrich2 as a possible novel Gln sensor that regulates microtubule glutamylation and mitochondrial function in mouse sperm.
Assuntos
Glutamina , Infertilidade Masculina , Animais , Humanos , Masculino , Camundongos , Ácido Glutâmico , Infertilidade Masculina/genética , Camundongos Knockout , Microtúbulos , Mitocôndrias , Proteínas Mitocondriais , Sêmen , Motilidade dos Espermatozoides , Espermatozoides , Tubulina (Proteína)RESUMO
Transcription factor-like 5 (TCFL5) is a testis-specific protein that contains the basic helix-loop-helix domain, but the in vivo functions of TCFL5 remain unknown. Herein, we generated CRISPR/Cas9-mediated knockout mice to dissect the function of TCFL5 in mouse testes. Surprisingly, we found that it was difficult to generate homozygous mice with the Tcfl5 deletion as the heterozygous males (Tcfl5+/-) were infertile. However, we did observe markedly abnormal phenotypes of spermatids and spermatozoa in the testes and epididymides of Tcfl5+/- mice. Mechanistically, we demonstrated that TCFL5 transcriptionally and post-transcriptionally regulated a set of genes participating in male germ cell development via TCFL5 ChIP-DNA and eCLIP-RNA high-throughput sequencing. We also identified a known RNA-binding protein, FXR1, as an interacting partner of TCFL5 that may coordinate the transition and localization of TCFL5 in the nucleus. Collectively, we herein report for the first time that Tcfl5 is haploinsufficient in vivo and acts as a dual-function protein that mediates DNA and RNA to regulate spermatogenesis. This article has an associated First Person interview with the first author of the paper.
Assuntos
Espermatogênese , Testículo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos , DNA/metabolismo , Fertilidade/genética , Masculino , Camundongos , Camundongos Knockout , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Espermatogênese/genética , Espermatozoides/metabolismo , Testículo/metabolismo , Fatores de Transcrição/metabolismoRESUMO
Mitochondria play a crucial role in spermatogenesis and are regulated by several mitochondrial fusion proteins. However, their functional importance associated with their structure formation and mRNA fate regulation during spermatogenesis remains unclear. Here, we show that mitofusin 2 (MFN2), a mitochondrial fusion protein, interacts with nuage-associated proteins (including MIWI, DDX4, TDRKH and GASZ) in mice. Conditional mutation of Mfn2 in postnatal germ cells results in male sterility due to germ cell developmental defects. Moreover, MFN2 interacts with MFN1, another mitochondrial fusion protein with a high-sequence similarity to MFN2, in testes to facilitate spermatogenesis. Simultaneous mutation of Mfn1 and Mfn2 in testes causes very severe infertile phenotypes. Importantly, we show that MFN2 is enriched in polysome fractions of testes and interacts with MSY2, a germ cell-specific DNA/RNA-binding protein, to control gamete-specific mRNA (such as Spata19) translational activity during spermatogenesis. Collectively, our findings demonstrate that MFN2 interacts with nuage-associated proteins and MSY2 to regulate male germ cell development by controlling several gamete-specific mRNA fates.
Assuntos
Diferenciação Celular/fisiologia , GTP Fosfo-Hidrolases/genética , GTP Fosfo-Hidrolases/metabolismo , Células Germinativas/metabolismo , RNA Mensageiro/metabolismo , Espermatogênese/genética , Espermatogênese/fisiologia , Animais , Proteínas Argonautas , RNA Helicases DEAD-box , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Fertilidade , Regulação da Expressão Gênica no Desenvolvimento , Técnicas de Inativação de Genes , Células Germinativas/patologia , Células HEK293 , Humanos , Infertilidade Masculina/genética , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Mitocôndrias/metabolismo , Dinâmica Mitocondrial , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Mutação , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Testículo/metabolismo , Testículo/patologiaRESUMO
Sertoli cells act as highly polarized testicular cells that nutritionally support multiple stages of germ cell development. However, the gene regulation network in Sertoli cells for modulating germ cell development has yet to be fully understood. In this study, we report that heterogeneous nuclear ribonucleoproteins C in Sertoli cells are essential for germ cell development and male fertility. Conditional knockout of heterogeneous nuclear ribonucleoprotein C in mouse Sertoli cells leads to aberrant Sertoli cells proliferation, disrupted cytoskeleton of Sertoli cells, and compromised blood-testis barrier function, resulting in loss of supportive cell function and, ultimately, defective spermiogenesis in mice. Further ribonucleic acid-sequencing analyses revealed these phenotypes are likely caused by the dysregulated genes in heterogeneous nuclear ribonucleoprotein C-deficient Sertoli cells related to cell adhesion, cell proliferation, and apoptotic process. In conclusion, this study demonstrates that heterogeneous nuclear ribonucleoprotein C plays a critical role in Sertoli cells for maintaining the function of Sertoli cells and sustaining steady-state spermatogenesis in mice.
Assuntos
Fertilidade , Camundongos Knockout , Células de Sertoli , Espermatogênese , Animais , Masculino , Células de Sertoli/metabolismo , Células de Sertoli/fisiologia , Espermatogênese/fisiologia , Espermatogênese/genética , Camundongos , Fertilidade/fisiologia , Fertilidade/genética , Ribonucleoproteínas Nucleares Heterogêneas Grupo C/metabolismo , Ribonucleoproteínas Nucleares Heterogêneas Grupo C/genética , Barreira Hematotesticular/metabolismoRESUMO
Mammalian oviducts play an essential role in female fertility by picking up ovulated oocytes and transporting and nurturing gametes (sperm/oocytes) and early embryos. However, the relative contributions to these functions from various cell types within the oviduct remain controversial. The oviduct in mice deficient in two microRNA (miRNA) clusters (miR-34b/c and miR-449) lacks cilia, thus allowing us to define the physiological role of oviductal motile cilia. Here, we report that the infundibulum without functional motile cilia failed to pick up the ovulated oocytes. In the absence of functional motile cilia, sperm could still reach the ampulla region, and early embryos managed to migrate to the uterus, but the efficiency was reduced. Further transcriptomic analyses revealed that the five messenger ribonucleic acids (mRNAs) encoded by miR-34b/c and miR-449 function to stabilize a large number of mRNAs involved in cilium organization and assembly and that Tubb4b was one of their target genes. Our data demonstrate that motile cilia in the infundibulum are essential for oocyte pickup and thus, female fertility, whereas motile cilia in other parts of the oviduct facilitate gamete and embryo transport but are not absolutely required for female fertility.
Assuntos
Cílios/fisiologia , Fertilidade , Oócitos/fisiologia , Oviductos/fisiologia , Ovulação , Animais , Blastocisto/fisiologia , Implantação do Embrião , Feminino , Masculino , Camundongos Knockout , MicroRNAs/metabolismo , Movimento , Espermatozoides/fisiologiaRESUMO
Mitochondria tailor their morphology to execute their specialized functions in different cell types and/or different environments. During spermatogenesis, mitochondria undergo continuous morphological and distributional changes with germ cell development. Deficiencies in these processes lead to mitochondrial dysfunction and abnormal spermatogenesis, thereby causing male infertility. In recent years, mitochondria have attracted considerable attention because of their unique role in the regulation of piRNA biogenesis in male germ cells. In this review, we describe the varied characters of mitochondria and focus on key mitochondrial factors that play pivotal roles in the regulation of spermatogenesis, from primordial germ cells to spermatozoa, especially concerning metabolic shift, stemness and reprogramming, mitochondrial transformation and rearrangement, and mitochondrial defects in human sperm. Further, we discuss the molecular mechanisms underlying these processes.
Assuntos
Diferenciação Celular , Células Germinativas/citologia , Infertilidade Masculina/patologia , Mitocôndrias/fisiologia , Doenças Mitocondriais/fisiopatologia , Espermatozoides/citologia , Animais , Humanos , Infertilidade Masculina/etiologia , Infertilidade Masculina/metabolismo , MasculinoRESUMO
The mouse WD repeat and FYVE domain containing 1 (Wdfy1) gene is located in chromosome 1qC4 and spans over 73.7 kilobases. It encodes a protein of 410-amino acid protein that shares 97.8% amino acid sequence identity with the human WDFY1 protein. However, the expression pattern of WDFY1 in reproductive organs and its function in male fertility remain unknown. In this study, we generated transgenic mice expressing FLAG-Wdfy1-mCherry cDNA driven by the Wdfy1 promoter to clarify the expression of WDFY1. The results showed that WDFY1 is highly expressed in mouse testes and located in the cytoplasm of late pachytene spermatocytes to elongated spermatids. Interestingly, the global Wdfy1 knockout (KO) male mice displayed normal growth, development, and fertility. Further histological analysis of Wdfy1 knockout mouse testes revealed that all spermatogenic cells are present in Wdfy1 KO seminiferous tubules. Together, our data demonstrate that WDFY1 is dispensable for mouse spermatogenesis and male fertility.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Fertilidade/genética , Regulação da Expressão Gênica , Espermatogênese/genética , Testículo/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Western Blotting , Feminino , Perfilação da Expressão Gênica/métodos , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Espermátides/citologia , Espermátides/metabolismo , Testículo/citologia , Repetições WD40/genéticaRESUMO
During male meiosis, the constitutively unsynapsed XY chromosomes undergo meiotic sex chromosome inactivation (MSCI), and the DNA damage response (DDR) pathway is critical for MSCI establishment. Our previous study showed that UHRF1 (ubiquitin-like, with PHD and ring finger domains 1) deletion led to meiotic arrest and male infertility; however, the underlying mechanisms of UHRF1 in the regulation of meiosis remain unclear. Here, we report that UHRF1 is required for MSCI and cooperates with the DDR pathway in male meiosis. UHRF1-deficient spermatocytes display aberrant pairing and synapsis of homologous chromosomes during the pachytene stage. In addition, UHRF1 deficiency leads to aberrant recruitment of ATR and FANCD2 on the sex chromosomes and disrupts the diffusion of ATR to the XY chromatin. Furthermore, we show that UHRF1 acts as a cofactor of BRCA1 to facilitate the recruitment of DDR factors onto sex chromosomes for MSCI establishment. Accordingly, deletion of UHRF1 leads to the failure of meiotic silencing on sex chromosomes, resulting in meiotic arrest. In addition to our previous findings, the present study reveals that UHRF1 participates in MSCI, ensuring the progression of male meiosis. This suggests a multifunctional role of UHRF1 in the male germline.
Assuntos
Proteínas Estimuladoras de Ligação a CCAAT , Pareamento Cromossômico , Cromossomos Sexuais , Ubiquitina-Proteína Ligases , Animais , Proteínas Estimuladoras de Ligação a CCAAT/genética , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Dano ao DNA , Masculino , Meiose/genética , Camundongos , Cromossomos Sexuais/genética , Espermatócitos/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismoRESUMO
Comment on "A microRNA cluster in the Fragile-X region expressed during spermatogenesis targets FMR1" by Ramaiah et al.
Assuntos
MicroRNAs , Animais , Proteína do X Frágil da Deficiência Intelectual , Regulação da Expressão Gênica , Masculino , Camundongos , Espermatogênese , EspermatogôniasRESUMO
Emerging evidence shows that m6A, one of the most abundant RNA modifications in mammals, is involved in the entire process of spermatogenesis, including mitosis, meiosis, and spermiogenesis. "Writers" catalyze m6A formation on stage-specific transcripts during male germline development, while "erasers" remove m6A modification to maintain a balance between methylation and demethylation. The different functions of RNA-m6A transcripts depend on their recognition by "readers". m6A modification mediates RNA metabolism, including mRNA splicing, translation, and degradation, as well as the maturity and biosynthesis of non-coding RNAs. Sperm RNA profiles are easily affected by environmental exposure and can even be inherited for several generations, similar to epigenetic inheritance. Here, we review and summarize the critical role of m6A in different developmental stages of male germ cells, to understand of the mechanisms and epigenetic regulation of m6A modifications. In addition, we also outline and discuss the important role of non-coding RNAs in spermatogenesis and RNA modifications in epigenetic inheritance.
Assuntos
Epigênese Genética , RNA/metabolismo , Espermatozoides/metabolismo , Enzimas AlkB/metabolismo , Animais , Humanos , Masculino , Metiltransferases/metabolismo , Fatores de Processamento de Serina-Arginina/metabolismo , EspermatogêneseRESUMO
Cilia are cell-surface, microtubule-based organelles that project into extracellular space. Motile cilia are conserved throughout eukaryotes, and their beat induces the flow of fluid, relative to cell surfaces. In mammals, the coordinated beat of motile cilia provides highly specialized functions associated with the movement of luminal contents, as seen with metachronal waves transporting mucus in the respiratory tract. Motile cilia are also present in the male and female reproductive tracts. In the female, wave-like motions of oviductal cilia transport oocytes and embryos toward the uterus. A similar function has been assumed for motile cilia in efferent ductules of the male-i.e., to transport immotile sperm from rete testis into the epididymis. However, we report here that efferent ductal cilia in the male do not display a uniform wave-like beat to transport sperm solely in one direction, but rather exert a centripetal force on luminal fluids through whip-like beating with continual changes in direction, generating turbulence, which maintains immotile spermatozoa in suspension within the lumen. Genetic ablation of two miRNA clusters (miR-34b/c and -449a/b/c) led to failure in multiciliogenesis in murine efferent ductules due to dysregulation of numerous genes, and this mouse model allowed us to demonstrate that loss of efferent duct motile cilia causes sperm aggregation and agglutination, luminal obstruction, and sperm granulomas, which, in turn, induce back-pressure atrophy of the testis and ultimately male infertility.
Assuntos
Cílios/genética , Infertilidade Masculina/genética , MicroRNAs/genética , Animais , Epididimo/crescimento & desenvolvimento , Epididimo/patologia , Feminino , Genitália Masculina/crescimento & desenvolvimento , Humanos , Infertilidade Masculina/fisiopatologia , Masculino , Camundongos , Camundongos Knockout , Microtúbulos/genética , Microtúbulos/metabolismo , Espermatozoides/crescimento & desenvolvimento , Espermatozoides/patologia , Testículo/crescimento & desenvolvimento , Testículo/metabolismoRESUMO
Protein methyltransferases play various physiological and pathological roles through methylating histone and non-histone targets. Many histone methyltransferases have been reported to regulate the development of spermatogenic cells. However, the specific function of non-histone methyltransferases during spermatogenesis remains unclear. In this study, we found that METTL21A, a non-histone methyltransferase, is highly expressed in mouse testes. In order to elucidate the role of METTL21A in spermatogenesis, we generated a Mettl21a global knockout mouse model using CRISPR/Cas9 technology. Unexpectedly, our results showed that knockout males are fertile without apparent defects in the processes of male germ cell development, including spermatogonial differentiation, meiosis, and sperm maturation. Furthermore, the ablation of METTL21A does not affect the expression and localization of its known targeting proteins in testes. Together, our data demonstrated that METTL21A is not essential for mouse spermatogenesis and male fertility.
Assuntos
Fertilidade/fisiologia , Histona Metiltransferases/metabolismo , Espermatogênese/fisiologia , Animais , Diferenciação Celular/fisiologia , Feminino , Masculino , Meiose/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Camundongos Knockout , Espermatogônias/metabolismo , Testículo/metabolismoRESUMO
Previous studies have shown that Dnmt2-null sperm block the paternal transmission (through sperm) of certain acquired traits, e.g., high-fat diet-induced metabolic disorders or white tails due to a Kit paramutation. Here, we report that DNMT2 is also required for the transmission of a Kit paramutant phenotype (white tail tip) through the female germline (i.e., oocytes). Specifically, ablation of Dnmt2 led to aberrant profiles of tRNA-derived small RNAs (tsRNAs) and other small noncoding RNAs (sncRNAs) in sperm, which correlate with altered mRNA transcriptomes in pronuclear zygotes derived from wild-type oocytes carrying the Kit paramutation and a complete blockage of transmission of the paramutant phenotype through oocytes. Together, the present study suggests that both paternal and maternal transmissions of epigenetic phenotypes require intact DNMT2 functions in the male germline.
Assuntos
DNA (Citosina-5-)-Metiltransferases/deficiência , Epigênese Genética , Camundongos/genética , Mutação , Pigmentação/genética , Proteínas Proto-Oncogênicas c-kit/genética , Cauda/fisiologia , Animais , Cor , DNA (Citosina-5-)-Metiltransferases/metabolismo , Feminino , Masculino , FenótipoRESUMO
Otogelin-like protein (encoded by Otogl) was highly structural similar to the gelforming mucin proteins. Although human OTOG mutations have been linked to deafness, the biological function of OTOGL in male germ cell development remains enigmatic. In screening 336 patients with non-obstructive azoospermia (NOA), OTOGL displays the high mutant ratio (13.99 %). Then, we examined the expression of OTOGL in developing mouse testes. Otogl mRNA and protein are continually expressed in postnatal developing testes from postnatal day 0 (P0) testes to P21 testes exhibiting a decreased trend with the age growth. We thus generated a global Otogl knockout mouse (KO) model using the CRISPR/Cas9 technology; however, Otogl KO mice displayed normal development and fertility. Further histological analysis of Otogl knockout mouse testes revealed that all types of spermatogenic cells are present in Otogl KO seminiferous tubules. Together, our study suggested that OTOGL is nonessential for male germ cell development and spermatogenesis.
Assuntos
Proteínas de Membrana/biossíntese , Mucinas/biossíntese , Espermatogênese/fisiologia , Testículo/metabolismo , Animais , Azoospermia/genética , Azoospermia/metabolismo , Azoospermia/patologia , Células Germinativas , Humanos , Masculino , Proteínas de Membrana/genética , Camundongos , Camundongos Knockout , Mucinas/genética , Mutação de Sentido Incorreto/fisiologia , Testículo/crescimento & desenvolvimentoRESUMO
Correct orientation of the mitotic spindle determines the plane of cellular cleavage and is crucial for organ development. In the developing cerebral cortex, spindle orientation defects result in severe neurodevelopmental disorders, but the precise mechanisms that control this important event are not fully understood. Here, we use a combination of high-content screening and mouse genetics to identify the miR-34/449 family as key regulators of mitotic spindle orientation in the developing cerebral cortex. By screening through all cortically expressed miRNAs in HeLa cells, we show that several members of the miR-34/449 family control mitotic duration and spindle rotation. Analysis of miR-34/449 knockout (KO) mouse embryos demonstrates significant spindle misorientation phenotypes in cortical progenitors, resulting in an excess of radial glia cells at the expense of intermediate progenitors and a significant delay in neurogenesis. We identify the junction adhesion molecule-A (JAM-A) as a key target for miR-34/449 in the developing cortex that might be responsible for those defects. Our data indicate that miRNA-dependent regulation of mitotic spindle orientation is crucial for cell fate specification during mammalian neurogenesis.
Assuntos
Córtex Cerebral/embriologia , MicroRNAs/metabolismo , Fuso Acromático/metabolismo , Animais , Células HeLa , Humanos , Camundongos , Camundongos KnockoutRESUMO
During acrosome biogenesis, numerous granules formed from trans-Golgi stacks and accumulated in the concave region of the nuclear surface that is essential for acrosome formation. Several Golgi-associated proteins were involved in this process. However, the specific function of Golgi-associated proteins, especially Golgi matrix protein, during acrosome biogenesis remains elusive. In this study, we identified GOLGA4, as a Golgi matrix protein, highly expressed in mouse testes. To explore the function of GOLGA4 in spermatogenesis, we generated a Golga4 global knockout mouse line using CRISPR/Cas9 technology and demonstrated that Golga4 knockout males are fertile with normal morphology of testis and sperm. Furthermore, testicular histology showed no significant difference between WT and KO mice. Together, our data demonstrate that GOLGA4 is dispensable for mouse spermatogenesis and male fertility.
Assuntos
Autoantígenos/genética , Fertilidade/genética , Perfilação da Expressão Gênica/métodos , Proteínas da Matriz do Complexo de Golgi/genética , Espermatogênese/genética , Animais , Autoantígenos/metabolismo , Sequência de Bases , Feminino , Proteínas da Matriz do Complexo de Golgi/metabolismo , Humanos , Fígado/metabolismo , Masculino , Camundongos Knockout , Ovário/metabolismo , Estômago/química , Testículo/metabolismoRESUMO
Of all human infertility cases, up to 50% show contributing factors leading to defects in the male reproductive physiology. Seminal plasma (SP) is the biological fluid derived from the male accessory sex gland which carries spermatozoa passing throughout the male and female reproductive tract during ejaculation. It contains a complicated set of heterogeneous molecular structures, including proteins, cell-free nucleic acid (DNA, microRNA and LncRNA), and small-molecule metabolites as well as inorganic chemicals (ions). For a long time, the substantial significance of seminal plasma factors' functions has been underestimated, which is restricted to spermatozoa transport and protection. Notably, significant advancements have been made in dissecting seminal plasma components, revealing new insights into multiple aspects of sperm function, as well as fertilization and pregnancy outcomes in recent years. In this review, we summarize the state-of-art discoveries regarding SP compositions and their implications in male fertility, particularly describing the novel understanding of seminal plasma components and related modifications using "omics" approaches and mainly focusing on proteome and RNA-seq data in the latest decade. Meanwhile, we highlighted the proposed mechanism of the regulation of SP molecules on immunomodulation in the female reproductive tract. Moreover, we also discussed the proteins investigated as non-invasive diagnosis biomarkers for male infertility in the clinic.