RESUMEN
Acephalic spermatozoa syndrome (ASS) is a severe teratospermia with decaudated, decapitated, and malformed sperm, resulting in male infertility. Nuclear envelope protein SUN5 localizes to the junction between the sperm head and tail. Mutations in the SUN5 gene have been identified most frequently (33-47%) in ASS cases, and its molecular mechanism of action is yet to be explored. In the present study, we generated Sun5 knockout mice, which presented the phenotype of ASS. Nuclear membrane protein LaminB1 and cytoskeletal GTPases Septin12 and Septin2 were identified as potential partners for interacting with SUN5 by immunoprecipitation-mass spectrometry in mouse testis. Further studies demonstrated that SUN5 connected the nucleus by interacting with LaminB1 and connected the proximal centriole by interacting with Septin12. The binding between SUN5 and Septin12 promoted their aggregation together in the sperm neck. The disruption of the LaminB1/SUN5/Septin12 complex by Sun5 deficiency caused separation of the Septin12-proximal centriole from the nucleus, leading to the breakage of the head-to-tail junction. Collectively, these data provide new insights into the pathogenesis of ASS caused by SUN5 deficiency.
Asunto(s)
Proteínas de la Membrana , Ratones Noqueados , Membrana Nuclear , Septinas , Cabeza del Espermatozoide , Cola del Espermatozoide , Animales , Humanos , Masculino , Ratones , Infertilidad Masculina/metabolismo , Infertilidad Masculina/genética , Lamina Tipo B/metabolismo , Lamina Tipo B/genética , Proteínas de la Membrana/metabolismo , Proteínas de la Membrana/genética , Membrana Nuclear/metabolismo , Septinas/metabolismo , Septinas/genética , Cabeza del Espermatozoide/metabolismo , Cabeza del Espermatozoide/patología , Cola del Espermatozoide/metabolismo , Espermatozoides/metabolismo , Teratozoospermia/metabolismo , Teratozoospermia/genéticaRESUMEN
Sperm heads contain not only the nucleus but also the acrosome which is a distinctive cap-like structure located anterior to the nucleus and is derived from the Golgi apparatus. The Golgi Associated RAB2 Interactors (GARINs; also known as FAM71) protein family shows predominant expression in the testis and all possess a RAB2-binding domain which confers binding affinity to RAB2, a small GTPase that is responsible for membrane transport and vesicle trafficking. Our previous study showed that GARIN1A and GARIN1B are important for acrosome biogenesis and that GARIN1B is indispensable for male fertility in mice. Here, we generated KO mice of other Garins, namely Garin2, Garin3, Garin4, Garin5a, and Garin5b (Garin2-5b). Using computer-assisted morphological analysis, we found that the loss of each Garin2-5b resulted in aberrant sperm head morphogenesis. While the fertilities of Garin2-/- and Garin4-/- males are normal, Garin5a-/- and Garin5b-/- males are subfertile, and Garin3-/- males are infertile. Further analysis revealed that Garin3-/- males exhibited abnormal acrosomal morphology, but not as severely as Garin1b-/- males; instead, the amounts of membrane proteins, particularly ADAM family proteins, decreased in Garin3 KO spermatozoa. Moreover, only Garin4 KO mice exhibit vacuoles in the sperm head. These results indicate that GARINs assure correct head morphogenesis and some members of the GARIN family function distinctively in male fertility.
Asunto(s)
Fertilidad , Infertilidad Masculina , Ratones Noqueados , Cabeza del Espermatozoide , Animales , Masculino , Ratones , Acrosoma/metabolismo , Fertilidad/genética , Aparato de Golgi/metabolismo , Infertilidad Masculina/genética , Infertilidad Masculina/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas de la Membrana/genética , Morfogénesis/genética , Proteína de Unión al GTP rab2/metabolismo , Proteína de Unión al GTP rab2/genética , Cabeza del Espermatozoide/metabolismo , Espermatozoides/metabolismo , Testículo/metabolismo , Testículo/crecimiento & desarrolloRESUMEN
The centrosome is critical for maintaining the sperm head-tail connection and the formation of flagellar microtubules. In this study, we found that in mouse testes, CCDC159 (coiled-coil domain-containing protein 159) is specifically localized to the head-tail coupling apparatus (HTCA) of spermatids, a structure that ensures sperm head-tail tight conjunction. CCDC159 contains a C-terminal coiled-coil domain that functions as the centrosomal localization signal. Gene knockout (KO) of Ccdc159 in mice resulted in acephalic spermatozoa, abnormal flagella, and male infertility. To explore the mechanism behind CCDC159 regulating spermatogenesis, we identified CCDC159-binding proteins using a yeast two-hybrid screen and speculated that CCDC159 participates in HTCA assembly by regulating protein phosphatase PP1 activity. Further RNA-sequencing analyses of Ccdc159 KO testes revealed numerous genes involved in male gamete generation that were downregulated. Together, our results show that CCDC159 in spermatids is a novel centrosomal protein anchoring the sperm head to the tail. Considering the limitation of KO mouse model in clarifying the biological function of CCDC159 in spermatogenesis, a gene-rescue experiment will be performed in the future.
Asunto(s)
Ratones Noqueados , Cabeza del Espermatozoide , Cola del Espermatozoide , Espermátides , Espermatogénesis , Animales , Masculino , Ratones , Espermátides/metabolismo , Cola del Espermatozoide/metabolismo , Espermatogénesis/fisiología , Cabeza del Espermatozoide/metabolismo , Infertilidad Masculina/genética , Infertilidad Masculina/metabolismo , Testículo/metabolismo , Centrosoma/metabolismoRESUMEN
Approximately 10%-15% of couples worldwide are infertile, and male factors account for approximately half of these cases. Teratozoospermia is a major cause of male infertility. Although various mutations have been identified in teratozoospermia, these can vary among ethnic groups. In this study, we performed whole-exome sequencing to identify genetic changes potentially causative of teratozoospermia. Out of seven genes identified, one, ATP/GTP Binding Protein 1 (AGTPBP1), was characterized, and three missense changes were identified in two patients (Affected A: p.Glu423Asp and p.Pro631Leu; Affected B: p.Arg811His). In those two cases, severe sperm head and tail defects were observed. Moreover, AGTPBP1 localization showed a fragmented pattern compared to control participants, with specific localization in the neck and annulus regions. Using murine models, we found that AGTPBP1 is localized in the manchette structure, which is essential for sperm structure formation. Additionally, in Agtpbp1-null mice, we observed sperm head and tail defects similar to those in sperm from AGTPBP1-mutated cases, along with abnormal polyglutamylation tubulin and decreasing â³-2 tubulin levels. In this study, we established a link between genetic changes in AGTPBP1 and human teratozoospermia for the first time and identified the role of AGTPBP1 in deglutamination, which is crucial for sperm formation.
Asunto(s)
Infertilidad Masculina , D-Ala-D-Ala Carboxipeptidasa de Tipo Serina , Teratozoospermia , Humanos , Masculino , Animales , Ratones , Teratozoospermia/genética , Teratozoospermia/metabolismo , Tubulina (Proteína)/metabolismo , Semen/metabolismo , Espermatozoides/metabolismo , Cabeza del Espermatozoide/metabolismo , Flagelos/metabolismo , Infertilidad Masculina/genética , Infertilidad Masculina/metabolismo , Mutación , Proteínas de Unión al GTP/metabolismo , D-Ala-D-Ala Carboxipeptidasa de Tipo Serina/genética , D-Ala-D-Ala Carboxipeptidasa de Tipo Serina/metabolismoRESUMEN
Sperm with normal morphology and motility are essential for successful fertilization, and the strong attachment of the sperm head-tail coupling apparatus to the nuclear envelope during spermatogenesis is required to ensure the integrity of sperm for capacitation and fertilization. Here, we report that Arrdc5 is associated with spermatogenesis. The Arrdc5 knockout mouse model showed male infertility characterized by a high bent-head rate and reduced motility in sperm, which led to capacitation defects and subsequent fertilization failure. Through mass spectrometry, we found that ARRDC5 affects spermatogenesis by affecting NDC1 and SUN5. We further found that ARRDC5 might affect the vesicle-trafficking protein SEC22A-mediated transport and localization of NDC1, SUN5 and other head-tail coupling apparatus-related proteins that are responsible for initiating the attachment of the sperm head and tail. We finally performed intracytoplasmic sperm injection as a way to explore therapeutic strategies. Our findings demonstrate the essential role and the underlying molecular mechanism of ARRDC5 in anchoring the sperm head to the tail during spermatogenesis.
Asunto(s)
Infertilidad Masculina , Semen , Humanos , Animales , Ratones , Masculino , Semen/metabolismo , Espermatogénesis/genética , Espermatozoides/metabolismo , Cabeza del Espermatozoide/metabolismo , Proteínas/metabolismo , Infertilidad Masculina/genética , Infertilidad Masculina/metabolismo , Ratones Noqueados , Proteínas de la Membrana/metabolismoRESUMEN
Acephalic spermatozoa syndrome (ASS) is a rare and severe type of teratozoospermia characterized by the predominance of headless spermatozoa in the ejaculate. However, knowledge about the causative genes associated with ASS in humans is limited. Loss-of-function of SPATA20 has been suggested to result in the separation of the sperm head and flagellum in mice, whereas there have been no cases reporting SPATA20 variants leading to human male infertility. In this study, a nonsense mutation in SPATA20 (c.619C > T, p.Arg207*) was first identified in an ASS patient. Moreover, this variant contributed to the degradation of SPATA20 and was associated with decreased expression of SPATA6, which plays a vital role in the assembly of the sperm head-tail conjunction in humans. In addition, the infertility caused by loss-of-function mutation of SPATA20 might not be rescued by intracytoplasmic sperm injection (ICSI). Collectively, our findings suggested that SPATA20 might be required for sperm head-tail conjunction formation in humans, the nonfunction of which may lead to male infertility related to ASS. The discovery of the loss-of-function mutation in SPATA20 enriches the gene variant spectrum of human ASS, further contributing to improved diagnosis, genetic counseling and prognosis for male infertility.
Asunto(s)
Infertilidad Masculina , Semen , Teratozoospermia , Humanos , Masculino , Proteínas del Citoesqueleto/genética , Infertilidad Masculina/genética , Mutación , Cabeza del Espermatozoide/metabolismo , Espermatozoides/metabolismo , Teratozoospermia/genéticaRESUMEN
ODF1 is a major protein of the accessory fibres of the mammalian sperm tail. In addition, ODF1 is found in the connecting piece, a complex structure located at the posterior end of the nucleus that connects the sperm head and tail. The tight coupling of the sperm head and tail is critical for the progressive motility of the sperm to reach the oocyte for fertilisation. The depletion of ODF1 by homologous recombination in mice led to male infertility. Although sperm tails were present in the epididymis, no intact spermatozoa were found. Instead, the depletion of ODF1 resulted in sperm decapitation, suggesting that ODF1 is essential for the formation of the coupling apparatus and the tight linkage of the sperm head and tail. However, the development of the linkage complex in the absence of ODF1 has never been investigated. Here, I analysed the fine structure of the developing connecting piece by transmission electron microscopy. I show that the connecting piece develops as in wild-type spermatids. Structural abnormalities were not observed when ODF1 was absent. Thus, ODF1 is dispensable for the development of the connecting piece. However, the decapitation of ODF1-deficient spermatozoa indicates that the heads and tails of the spermatozoa are not linked, so that they separate when force is applied.
Asunto(s)
Decapitación , Espermátides , Animales , Decapitación/metabolismo , Masculino , Mamíferos , Ratones , Semen , Cabeza del Espermatozoide/metabolismo , Cola del Espermatozoide/metabolismo , Espermatozoides/metabolismoRESUMEN
Nonobstructive azoospermia (NOA) is the most serious form of spermatogenesis abnormalities in male infertility. Genetic factors are important to consider as elements leading to NOA. Although many pathogenic genes have been reported, the causative genes of NOA for many patients are still unknown. In this study, we found ten point mutations in the gene encoding homeodomain-interacting protein kinase 4 (HIPK4) in patients with NOA, and using in vitro studies, we determined a premature termination point mutation (p. Lys490∗, c.1468A>T) that can cause decreased expression of HIPK4. Our phosphoproteomic analysis of Hipk4-/- testes revealed phosphorylation of multiple proteins regulated by HIPK4 during spermiogenesis. We also confirmed that a substrate of HIPK4 with four downregulated phosphorylation sites matching the xSPx motif is the known manchette-related protein RIMS-binding protein 3, which is required for sperm head morphogenesis. Therefore, we conclude HIPK4 regulates the phosphorylation of manchette protein RIMS-binding protein 3 and plays essential roles in sperm head shaping and male fertility.
Asunto(s)
Azoospermia , Codón sin Sentido , Proteínas del Citoesqueleto , Proteínas Serina-Treonina Quinasas , Cabeza del Espermatozoide , Espermatogénesis , Azoospermia/genética , Azoospermia/metabolismo , Proteínas del Citoesqueleto/metabolismo , Humanos , Masculino , Fosforilación , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Cabeza del Espermatozoide/metabolismo , Espermatogénesis/genética , Testículo/metabolismoRESUMEN
The perinuclear theca (PT) is a cytoskeletal element encapsulating the sperm nucleus; however, our understanding of the physiological roles of PT in sperm is very limited. We show that Calicin interacts with itself and many other PT components, indicating it may serve as an organizing center of the PT assembly. Calicin is detectable first when surrounding the acrosome, then detected around the entire nucleus, and finally translocated to the postacrosomal region of spermatid heads. Intriguingly, loss of Calicin specifically causes surface subsidence of sperm heads in the nuclear condensation stage. Calicin interacts with inner acrosomal membrane (IAM) protein Spaca1 and nuclear envelope (NE) components to form an "IAM-PT-NE" structure. Intriguingly, Ccin-knockout sperm also exhibit DNA damage and failure of fertilization. Our study provides solid animal evidence to suggest that the PT encapsulating sperm nucleus helps shape the sperm head and maintain the nuclear structure.
Asunto(s)
Proteínas del Citoesqueleto , Semen , Cabeza del Espermatozoide , Animales , Proteínas del Citoesqueleto/metabolismo , Citoesqueleto/fisiología , Masculino , Ratones , Semen/metabolismo , Cabeza del Espermatozoide/metabolismo , Cabeza del Espermatozoide/fisiologíaRESUMEN
Alzheimer's is characterized by accumulation of amyloid-ß (Aß) associated with insufficient clearance of toxicants from the brain establishing a chronic inflammation and other abnormalities in the brain. Inflammatory microglia and astrocytes along with abnormal lymphatics associated with insufficient clearance of Aß and other toxicants from the brain establish a chronic inflammation. This causes abnormal choroid plexus, leukocyte trafficking, and hypoxic condition along with high levels of regulatory T cells (Tregs). There is no consensus among researchers regarding decreasing or increasing Tregs to achieve therapeutic effects. Different opposing studies tried to suppress or boost inflammation to treat AD. Based on reproductive immunology, sperm induces constructive inflammatory response and seminal-vesicle-fluid (SVF) suppresses inflammation leading to uterus remodeling. It prompted us to compare therapeutic efficiency of inflammatory or anti-inflammatory approaches in AD model based on reproductive immunology. To do so, SVF, sperm, or sperm head (from Wistar rat) was administered via intra-cerebro-ventricular route to Sprague Dawley rat AD model. Behavioral and histological examination were made and treatment groups were compared with control AD model and normal groups. Therapeutic efficacy was in the order of sperm head>sperm>SVF. Sperm head returned learning memory, Aß, lymphatics, neural growth factors, choroid plexus function, Iba-1/GFAP, MHC II/CD86/CD40, CD38/IL-10, and hypoxia levels back to normal level. However, SVF just partially ameliorated the disease. Immunologic properties of sperm/sperm head to elicit constructive inflammation can be extended to organs other than reproductive. This nature-based approach overcomes genetic difference as an important obstacle and limitation in cell therapy, and is expected to be safe or with least side effects.
Asunto(s)
Enfermedad de Alzheimer , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/terapia , Péptidos beta-Amiloides , Animales , Antiinflamatorios/uso terapéutico , Encéfalo , Tratamiento Basado en Trasplante de Células y Tejidos , Modelos Animales de Enfermedad , Femenino , Inflamación/metabolismo , Masculino , Ratas , Ratas Sprague-Dawley , Ratas Wistar , Cabeza del Espermatozoide/metabolismo , Cabeza del Espermatozoide/patologíaRESUMEN
The acrosome is a cap-shaped, Golgi-derived membranous organelle that is located over the anterior of the sperm nucleus and highly conserved throughout evolution. Although morphological changes during acrosome biogenesis in spermatogenesis have been well described, the molecular mechanism underlying this process is still largely unknown. Family with sequence similarity 71, member F1 and F2 (FAM71F1 and FAM71F2) are testis-enriched proteins that contain a RAB2B-binding domain, a small GTPase involved in vesicle transport and membrane trafficking. Here, by generating mutant mice for each gene, we found that Fam71f1 is essential for male fertility. In Fam71f1-mutant mice, the acrosome was abnormally expanded at the round spermatid stage, likely because of enhanced vesicle trafficking. Mass spectrometry analysis after immunoprecipitation indicated that, in testes, FAM71F1 binds not only RAB2B, but also RAB2A. Further study suggested that FAM71F1 binds to the GTP-bound active form of RAB2A/B, but not the inactive form. These results indicate that a complex of FAM71F1 and active RAB2A/B suppresses excessive vesicle trafficking during acrosome formation.
Asunto(s)
Acrosoma/metabolismo , Fertilidad/fisiología , Proteínas Nucleares/metabolismo , Proteínas de Unión al GTP rab/metabolismo , Proteína de Unión al GTP rab2/metabolismo , Acrosoma/patología , Animales , Genética , Aparato de Golgi/metabolismo , Infertilidad Masculina , Masculino , Ratones , Ratones Transgénicos , Proteínas Nucleares/genética , Unión Proteica , Cabeza del Espermatozoide/metabolismo , Espermatogénesis , Teratozoospermia/metabolismo , Testículo/metabolismoRESUMEN
Capacitation begins in the sperm head plasma membrane (HPM). Membrane rafts could house signaling molecules, but although these specialized microdomains have been microscopically visualized in sperm heads, rafts have been isolated for study only from homogenized whole sperm or tails, never purified HPM. Sodium/potassium ATPase (Na+ K+ -ATPase) is a membrane-bound signaling protein that induces capacitation in bull sperm in response to the steroid hormone ouabain, and its subunit isoforms α1, α3, ß1, ß2, and ß3 are known in HPM. This study hypothesized that rafts exist in the HPM of bull sperm, with Na+ K+ -ATPase subunit isoforms preferentially localized there. Western immunoblotting (WB) of HPM from fresh, uncapacitated bull sperm (n = 7 ejaculates), and detergent-resistant membranes isolated by density gradient centrifugation from this HPM, contained the raft-marker protein Flotillin-1; the non-raft fraction did not. HPM, raft, and non-raft contained all known Na+ K+ -ATPase isoforms including, for the first time, the previously unknown α2 isoform. Quantification (ImageQuant Software) found α3 and ß1 were relatively dominant isoforms in the HPM raft. WB profiles of raft isoforms differed significantly from HPM and non-raft profiles, with unique banding patterns and amounts, hinting that the capacitation signaling in the now-identified HPM rafts may depend on unique sequences within the isoform structure.
Asunto(s)
Ouabaína , ATPasa Intercambiadora de Sodio-Potasio , Animales , Bovinos , Masculino , Microdominios de Membrana/metabolismo , Ouabaína/metabolismo , Ouabaína/farmacología , Isoformas de Proteínas/metabolismo , Sodio , ATPasa Intercambiadora de Sodio-Potasio/metabolismo , Cabeza del Espermatozoide/metabolismoRESUMEN
The sperm head-to-tail coupling apparatus (HTCA) ensures sperm head-tail integrity while defective HTCA causes acephalic spermatozoa, rendering males infertile. Here, we show that CENTLEIN is indispensable for HTCA integrity and function, and that inactivation of CENTLEIN in mice leads to sperm decapitation and male sterility. We demonstrate that CENTLEIN directly interacts with both SUN5 and PMFBP1, two proteins localized in the HTCA and related with acephalic spermatozoa syndrome. We find that the absence of Centlein sets SUN5 and PMFBP1 apart, the former close to the sperm head and the latter in the decapitated tail. We show that lack of Sun5 results in CENTLEIN and PMFBP1 left in the decapitated tail, while disruption of Pmfbp1 results in SUN5 and CENTLEIN left on the detached sperm head. These results demonstrate that CENTLEIN cooperating with SUN5 and PMFBP1 participates in the HTCA assembly and integration of sperm head to the tail, indicating that impairments of CENTLEIN might be associated with acephalic spermatozoa syndrome in humans.
Asunto(s)
Proteínas del Citoesqueleto/metabolismo , Proteínas de la Membrana/metabolismo , Cabeza del Espermatozoide/metabolismo , Cola del Espermatozoide/metabolismo , Espermatozoides/metabolismo , Animales , Proteínas de Ciclo Celular , Células Cultivadas , Proteínas del Citoesqueleto/genética , Células HEK293 , Humanos , Infertilidad Masculina/genética , Infertilidad Masculina/metabolismo , Masculino , Proteínas de la Membrana/genética , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Ratones Noqueados , Mutación , Unión Proteica , Espermatozoides/citología , Teratozoospermia/genética , Teratozoospermia/metabolismoRESUMEN
The perinuclear theca (PT) of the eutherian sperm head is a cytoskeletal-like structure that houses proteins involved in important cellular processes during spermiogenesis and fertilization. Building upon our novel discovery of non-nuclear histones in the bovine PT, we sought to investigate whether this PT localization was a conserved feature of eutherian sperm. Employing cell fractionation, immunodetection, mass spectrometry, qPCR, and intracytoplasmic sperm injections (ICSI), we examined the localization, developmental origin, and functional potential of histones from the murid PT. Immunodetection localized histones to the post-acrosomal sheath (PAS) and the perforatorium (PERF) of the PT but showed an absence in the sperm nucleus. MS/MS analysis of selectively extracted PT histones indicated that predominately core histones (i.e., H3, H3.3, H2B, H2A, H2AX, and H4) populate the murid PT. These core histones appear to be de novo-synthesized in round spermatids and assembled via the manchette during spermatid elongation. Mouse ICSI results suggest that early embryonic development is delayed in the absence of PT-derived core histones. Here, we provide evidence that core histones are de novo-synthesized prior to PT assembly and deposited in PT sub-compartments for subsequent involvement in chromatin remodeling of the male pronucleus post-fertilization.
Asunto(s)
Histonas/biosíntesis , Cabeza del Espermatozoide/metabolismo , Espermátides/metabolismo , Espermatogénesis/fisiología , Animales , Núcleo Celular/metabolismo , Cromatografía Liquida/métodos , Femenino , Fertilización/fisiología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratas , Ratas Sprague-Dawley , Inyecciones de Esperma Intracitoplasmáticas , Espectrometría de Masas en Tándem/métodosRESUMEN
Infertility is a problem affecting an increasing number of couples worldwide. Currently, marker tests for male factor infertility are complex, highly technical and relatively subjective. Up to 40% of cases of male factor infertility are currently diagnosed as idiopathic therefore, there is a clear need for further research into better ways of diagnosing it. Changes in sperm telomere length have been associated with infertility and closely linked to DNA damage and fragmentation, which are also known to be related to infertility. However, telomere distribution is a parameter thus far underexplored as an infertility marker. Here, we assessed morphological parameters of sperm nuclei in fertile control and male factor infertile cohorts. In addition, we used 2D and 3D fluorescence in situ hybridization (FISH) to compare telomere distribution between these two groups. Our findings indicate that the infertile cohort sperm nuclei were, on average, 2.9% larger in area and showed subtle differences in sperm head height and width. Telomeres were mainly distributed towards the periphery of the nuclei in the control cohort, with diminishing telomere signals towards the center of the nuclei. Sperm nuclei of infertile males, however, had more telomere signals towards the center of the nuclei, a finding supported by 3D imaging. We conclude that, with further development, both morphology and telomere distribution may prove useful investigative tools in the fertility clinic.
Asunto(s)
Daño del ADN , Infertilidad Masculina/metabolismo , Infertilidad Masculina/patología , Cabeza del Espermatozoide/patología , Telómero/metabolismo , Biomarcadores/metabolismo , Estudios de Casos y Controles , Núcleo Celular/genética , Núcleo Celular/metabolismo , Núcleo Celular/patología , Estudios de Cohortes , Humanos , Infertilidad Masculina/genética , Masculino , Cabeza del Espermatozoide/metabolismo , Telómero/genéticaRESUMEN
Asthenoteratospermia is a common cause of male infertility. Recent studies have revealed that CFAP65 mutations lead to severe asthenoteratospermia due to acrosome hypoplasia and flagellum malformations. However, the molecular mechanism underlying CFAP65-associated sperm malformation is largely unclear. Here, we initially examined the role of CFAP65 during spermiogenesis using Cfap65 knockout (Cfap65-/-) mice. The results showed that Cfap65-/- male mice exhibited severe asthenoteratospermia characterized by morphologically defective sperm heads and flagella. In Cfap65-/- mouse testes, hyper-constricted sperm heads were apparent in step 9 spermatids accompanied by abnormal manchette development, and acrosome biogenesis was abnormal in the maturation phase. Moreover, subsequent flagellar elongation was also severely affected and characterized by disrupted assembly of the mitochondrial sheath (MS) in Cfap65-/- male mice. Furthermore, the proteomic analysis revealed that the proteostatic system during acrosome formation, manchette organization and MS assembly was disrupted when CFAP65 was lost. Importantly, endogenous immunoprecipitation and immunostaining experiments revealed that CFAP65 may form a cytoplasmic protein network comprising MNS1, RSPH1, TPPP2, ZPBP1 and SPACA1. Overall, these findings provide insights into the complex molecular mechanisms of spermiogenesis by uncovering the essential roles of CFAP65 during sperm head shaping, acrosome biogenesis and MS assembly.
Asunto(s)
Acrosoma/metabolismo , Proteínas de la Membrana/genética , Mitocondrias/genética , Mitocondrias/metabolismo , Espermatogénesis , Animales , Flagelos/genética , Flagelos/metabolismo , Flagelos/patología , Inmunohistoquímica , Infertilidad Masculina/genética , Masculino , Proteínas de la Membrana/metabolismo , Ratones , Ratones Noqueados , Mitocondrias/ultraestructura , Mapeo de Interacción de Proteínas , Mapas de Interacción de Proteínas , Cabeza del Espermatozoide/metabolismo , Cabeza del Espermatozoide/patología , Cola del Espermatozoide/metabolismo , Cola del Espermatozoide/patología , Cola del Espermatozoide/ultraestructura , Espermatogénesis/genética , Testículo/metabolismo , Testículo/patologíaRESUMEN
Spermatozoa are polarized cells with a head and a flagellum joined by the connecting piece. Head integrity is critical for normal sperm function, and head defects consistently lead to male infertility. Abnormalities of the sperm head are among the most severe and characteristic sperm defects. Patients presenting with a monomorphic head sperm defects such as globozoospermia or marcrozoospermia were analyzed permitting to identify several key genes for spermatogenesis such as AURKC and DPY19L2. The study of patients with other specific sperm head defects such as acephalic spermatozoa have also enabled the identification of new infertility genes such as SUN5. Here, we review the genetic causes leading to morphological defects of sperm head. Advances in the genetics of male infertility are necessary to improve the management of infertility and will pave the road towards future strategies of treatments, especially for patients with the most severe phenotype as sperm head defects.
Asunto(s)
Cabeza del Espermatozoide/patología , Espermatozoides/anomalías , Teratozoospermia/genética , Aurora Quinasa C/genética , Humanos , Infertilidad Masculina/genética , Infertilidad Masculina/patología , Masculino , Proteínas de la Membrana/genética , Cabeza del Espermatozoide/metabolismo , Espermatogénesis/genética , Espermatozoides/metabolismo , Espermatozoides/patología , Teratozoospermia/patologíaRESUMEN
Development of spermatozoa requires remodelling and formation of particular structures. In elongating spermatids, the transient microtubular manchette contributes to the formation of the head-tail coupling apparatus (HTCA) and the sperm tail. The HTCA derives from the centrosome in that the proximal centriole inserts into the nuclear indentation and the distal centriole gives rise to the sperm flagellum. Although impairments in the formation of HTCA and sperm tail cause male infertility their molecular constituents are only partially known. The WD40-protein CFAP52 is implicated in motile cilia, but its relevance for male germ cell differentiation is not known. Here we show that CFAP52 is widespread expressed and localizes to a subset of microtubular structures. In male germ cells, CFAP52 is a component of the transient manchette and the sperm tail. However, expression of Cfap52 is not restricted to motile cilia-bearing cells. In NIH3T3 cells, CFAP52 localizes to the centrosome, the basal body, and the mitotic spindle poles, but not to the primary cilium. Our results demonstrate that CFAP52 is not restricted to motile cilia but instead most likely functions in constituting the centrosome/basal body matrix and the sperm tail.
Asunto(s)
Proteínas Portadoras/metabolismo , Cilios/metabolismo , Flagelos/metabolismo , Espermátides/metabolismo , Animales , Cuerpos Basales/metabolismo , Proteínas Portadoras/genética , Centriolos/metabolismo , Centrosoma/metabolismo , Cilios/genética , Citoesqueleto/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Microtúbulos/metabolismo , Células 3T3 NIH , Cabeza del Espermatozoide/metabolismo , Cola del Espermatozoide/metabolismo , Espermatogénesis/fisiologíaRESUMEN
Sperm head shaping is a key event in spermiogenesis and is tightly controlled via the acrosome-manchette network. Linker of nucleoskeleton and cytoskeleton (LINC) complexes consist of Sad1 and UNC84 domain-containing (SUN) and Klarsicht/ANC-1/Syne-1 homology (KASH) domain proteins and form conserved nuclear envelope bridges implicated in transducing mechanical forces from the manchette to sculpt sperm nuclei into a hook-like shape. However, the role of LINC complexes in sperm head shaping is still poorly understood. Here we assessed the role of SUN3, a testis-specific LINC component harboring a conserved SUN domain, in spermiogenesis. We show that CRISPR/Cas9-generated Sun3 knockout male mice are infertile, displaying drastically reduced sperm counts and a globozoospermia-like phenotype, including a missing, mislocalized, or fragmented acrosome, as well as multiple defects in sperm flagella. Further examination revealed that the sperm head abnormalities are apparent at step 9 and that the sperm nuclei fail to elongate because of the absence of manchette microtubules and perinuclear rings. These observations indicate that Sun3 deletion likely impairs the ability of the LINC complex to transduce the cytoskeletal force to the nuclear envelope, required for sperm head elongation. We also found that SUN3 interacts with SUN4 in mouse testes and that the level of SUN4 proteins is drastically reduced in Sun3-null mice. Altogether, our results indicate that SUN3 is essential for sperm head shaping and male fertility, providing molecular clues regarding the underlying pathology of the globozoospermia-like phenotype.
Asunto(s)
Núcleo Celular/metabolismo , Forma de la Célula , Proteínas Nucleares/metabolismo , Cabeza del Espermatozoide/metabolismo , Espermatogénesis , Acrosoma/metabolismo , Animales , Núcleo Celular/genética , Eliminación de Gen , Masculino , Ratones , Ratones Noqueados , Proteínas Nucleares/genéticaRESUMEN
The centriole, or basal body, is the center of attachment between the sperm head and tail. While the distal end of the centriole templates the cilia, the proximal end associates with the nucleus. Using Drosophila, we identify a centriole-centric mechanism that ensures proper proximal end docking to the nucleus. This mechanism relies on the restriction of pericentrin-like protein (PLP) and the pericentriolar material (PCM) to the proximal end of the centriole. PLP is restricted proximally by limiting its mRNA and protein to the earliest stages of centriole elongation. Ectopic positioning of PLP to more distal portions of the centriole is sufficient to redistribute PCM and microtubules along the entire centriole length. This results in erroneous, lateral centriole docking to the nucleus, leading to spermatid decapitation as a result of a failure to form a stable head-tail linkage.