Aim： To show whether molecular motor dynein on a microtubule track, molecular motor myosin Va, motor recruiter myosin Va, VIIa-Rab27a/b interacting protein （MyRIP）, and vesicle receptor Rab27b on an F-actin track w...Aim： To show whether molecular motor dynein on a microtubule track, molecular motor myosin Va, motor recruiter myosin Va, VIIa-Rab27a/b interacting protein （MyRIP）, and vesicle receptor Rab27b on an F-actin track were present during human and monkey spermiogenesis involving intramanchette transport （IMT）. Methods： Spermiogenic cells were obtained from three men with obstructive azoospermia and normal adult cynomolgus monkey （Macacafascieularis）. Immunocytochemical detection and reverse transcription-polymerase chain reaction （RT-PCR） analysis of the proteins were carried out. Samples were analyzed by light microscope. Results： Using RT-PCR, we found that dynein, myosin Va, MyRIP and Rab27b were expressed in monkey testis. These proteins were localized to the manchette, as shown by immunofluorescence, particularly during human and monkey spermiogenesis. Conclusion： We speculate that during primate spermiogenesis, those proteins that compose microtubule-based and actin-based vesicle transport systems are actually present in the manchette and might possibly be involved in intramanchette transport.展开更多
Asthenoteratozoospermia is a common cause of male infertility.To further define the genetic causes underlying asthenoteratozoospermia,we performed whole-exome sequencing in a cohort of Han Chinese men with asthenotera...Asthenoteratozoospermia is a common cause of male infertility.To further define the genetic causes underlying asthenoteratozoospermia,we performed whole-exome sequencing in a cohort of Han Chinese men with asthenoteratozoospermia.Homozygous deleterious variants of MYCBPAP were first identified in two unrelated Chinese cases.Replication analyses in a French cohort revealed an additional asthenoteratozoospermia-affected case harboring a homozygous nonsense variant in MYCBPAP.All of the identified MYCBPAP variants were absent or extremely rare in the public human genome databases.Further functional assays indicated remarkably reduced abundance of MYCBPAP in the spermatozoa from MYCBPAP-associated cases.Subsequently,we generated a Mycbpap knockout(Mycbpap^(−/−))mouse model,which also exhibited male infertility with reduced sperm motility and abnormal morphologies in sperm heads and flagella.Further investigations demonstrated that Mycbpap^(−/−)male mice presented disrupted acrosome biogenesis and abnormally elongated manchette during spermiogenesis.Intriguingly,proteomic analyses indicated that the proteins related to spermatogenesis,acrosomal and flagellar functions were significantly down-regulated in the testes from Mycbpap^(−/−)male mice.Endogenous immunoprecipitation combined with mass spectrometry revealed interactions of MYCBPAP with a ribosome elimination related protein ARMC3 and central apparatus proteins including CFAP65 and CFAP70.Furthermore,MYCBPAP-associated male infertility in humans and mice could be partially overcome by using intracytoplasmic sperm injections.Collectively,these findings illustrate the essential role of MYCBPAP in normal spermatogenesis and homozygous deleterious variants in MYCBPAP can be considered as a genetic diagnostic indicator for infertile men with asthenoteratozoospermia.Our study will provide effective guidance for genetic counseling,clinical diagnosis and assisted reproduction treatments of MYCBPAP-associated male infertility.展开更多
Mammalian spermatogenesis is a well-organized process of cell development and differentiation. Meiosis expressed gene I （MEIG 1） plays an essential role in the regulation of spermiogenesis. To explore potential mech...Mammalian spermatogenesis is a well-organized process of cell development and differentiation. Meiosis expressed gene I （MEIG 1） plays an essential role in the regulation of spermiogenesis. To explore potential mechanisms of MEIGI＇s action, a yeast two-hybrid screen was conducted, and several potential binding partners were identified; one of them was membrane occupation and recognition nexus repeat containing 3 （MORN3）. MORN3 mRNA is only abundant in mouse testis. In the testis, Morn3 mRNA is highly expressed in the spermiogenesis stage. Specific anti-MORN3 polyclonal antibody was generated against N-terminus of the full-length MORN3 protein, and MORN3 expression and localization was examined in vitro and in vivo. In transfected Chinese hamster ovary cells, the antibody specifically crossed-reacted the full-length MORN3 protein, and immunofluorescence staining revealed that MORN3 was localized throughout the cytoplasm. Among multiple mouse tissues, about 25 kDa protein, was identified only in the testis. The protein was highly expressed after day 20 of birth. Immunofluorescence staining on mixed testicular cells isolated from adult wild-type mice demonstrated that MORN3 was expressed in the acrosome in germ cells throughout spermiogenesis. The protein was also present in the manchette of elongating spermatids. The total MORN3 expression and acrosome localization were not changed in the Meig/-deficient mice. However, its expression in manchette was dramatically reduced in the mutant mice. Our studies suggest that MORN3 is another regulator for spermatogenesis, probably together with MEIG1.展开更多
文摘Aim： To show whether molecular motor dynein on a microtubule track, molecular motor myosin Va, motor recruiter myosin Va, VIIa-Rab27a/b interacting protein （MyRIP）, and vesicle receptor Rab27b on an F-actin track were present during human and monkey spermiogenesis involving intramanchette transport （IMT）. Methods： Spermiogenic cells were obtained from three men with obstructive azoospermia and normal adult cynomolgus monkey （Macacafascieularis）. Immunocytochemical detection and reverse transcription-polymerase chain reaction （RT-PCR） analysis of the proteins were carried out. Samples were analyzed by light microscope. Results： Using RT-PCR, we found that dynein, myosin Va, MyRIP and Rab27b were expressed in monkey testis. These proteins were localized to the manchette, as shown by immunofluorescence, particularly during human and monkey spermiogenesis. Conclusion： We speculate that during primate spermiogenesis, those proteins that compose microtubule-based and actin-based vesicle transport systems are actually present in the manchette and might possibly be involved in intramanchette transport.
基金supported by the National Natural Science Foundation of China(32288101,32100480,32370654,82271638,32322017)the Innovative Research Team of High-Level Local Universities in Shanghai(SHSMU-ZDCX20212200)+1 种基金Shanghai Hospital Development Center Foundation(SHDC12023121)the outstanding Youth Foundation of Hunan Provincial Natural Science Foundation of China(2023JJ20080).
文摘Asthenoteratozoospermia is a common cause of male infertility.To further define the genetic causes underlying asthenoteratozoospermia,we performed whole-exome sequencing in a cohort of Han Chinese men with asthenoteratozoospermia.Homozygous deleterious variants of MYCBPAP were first identified in two unrelated Chinese cases.Replication analyses in a French cohort revealed an additional asthenoteratozoospermia-affected case harboring a homozygous nonsense variant in MYCBPAP.All of the identified MYCBPAP variants were absent or extremely rare in the public human genome databases.Further functional assays indicated remarkably reduced abundance of MYCBPAP in the spermatozoa from MYCBPAP-associated cases.Subsequently,we generated a Mycbpap knockout(Mycbpap^(−/−))mouse model,which also exhibited male infertility with reduced sperm motility and abnormal morphologies in sperm heads and flagella.Further investigations demonstrated that Mycbpap^(−/−)male mice presented disrupted acrosome biogenesis and abnormally elongated manchette during spermiogenesis.Intriguingly,proteomic analyses indicated that the proteins related to spermatogenesis,acrosomal and flagellar functions were significantly down-regulated in the testes from Mycbpap^(−/−)male mice.Endogenous immunoprecipitation combined with mass spectrometry revealed interactions of MYCBPAP with a ribosome elimination related protein ARMC3 and central apparatus proteins including CFAP65 and CFAP70.Furthermore,MYCBPAP-associated male infertility in humans and mice could be partially overcome by using intracytoplasmic sperm injections.Collectively,these findings illustrate the essential role of MYCBPAP in normal spermatogenesis and homozygous deleterious variants in MYCBPAP can be considered as a genetic diagnostic indicator for infertile men with asthenoteratozoospermia.Our study will provide effective guidance for genetic counseling,clinical diagnosis and assisted reproduction treatments of MYCBPAP-associated male infertility.
文摘Mammalian spermatogenesis is a well-organized process of cell development and differentiation. Meiosis expressed gene I （MEIG 1） plays an essential role in the regulation of spermiogenesis. To explore potential mechanisms of MEIGI＇s action, a yeast two-hybrid screen was conducted, and several potential binding partners were identified; one of them was membrane occupation and recognition nexus repeat containing 3 （MORN3）. MORN3 mRNA is only abundant in mouse testis. In the testis, Morn3 mRNA is highly expressed in the spermiogenesis stage. Specific anti-MORN3 polyclonal antibody was generated against N-terminus of the full-length MORN3 protein, and MORN3 expression and localization was examined in vitro and in vivo. In transfected Chinese hamster ovary cells, the antibody specifically crossed-reacted the full-length MORN3 protein, and immunofluorescence staining revealed that MORN3 was localized throughout the cytoplasm. Among multiple mouse tissues, about 25 kDa protein, was identified only in the testis. The protein was highly expressed after day 20 of birth. Immunofluorescence staining on mixed testicular cells isolated from adult wild-type mice demonstrated that MORN3 was expressed in the acrosome in germ cells throughout spermiogenesis. The protein was also present in the manchette of elongating spermatids. The total MORN3 expression and acrosome localization were not changed in the Meig/-deficient mice. However, its expression in manchette was dramatically reduced in the mutant mice. Our studies suggest that MORN3 is another regulator for spermatogenesis, probably together with MEIG1.
