Multiple morphological abnormalities of the flagella(MMAF)represent a severe form of sperm defects leading to asthenozoospermia and male infertility.In this study,we identified a novel homozygous splicing mutation(c.8...Multiple morphological abnormalities of the flagella(MMAF)represent a severe form of sperm defects leading to asthenozoospermia and male infertility.In this study,we identified a novel homozygous splicing mutation(c.871-4 ACA>A)in the adenylate kinase 7(AK7)gene by whole-exome sequencing in infertile individuals.Spermatozoa from affected individuals exhibited typical MMAF characteristics,including coiled,bent,short,absent,and irregular flagella.Transmission electron microscopy analysis showed disorganized axonemal structure and abnormal mitochondrial sheets in sperm flagella.Immunofluorescence staining confirmed the absence of AK7 protein from the patients’spermatozoa,validating the pathogenic nature of the mutation.This study provides direct evidence linking the AK7 gene to MMAF-associated asthenozoospermia in humans,expanding the mutational spectrum of AK7 and enhancing our understanding of the genetic basis of male infertility.展开更多
Multiple morphological abnormalities of the sperm flagella(MMAF)are characterized by bent,irregular,short,coiled,and absent flagella.MMAF is caused by a variety of genes,some of which have been identified.However,the ...Multiple morphological abnormalities of the sperm flagella(MMAF)are characterized by bent,irregular,short,coiled,and absent flagella.MMAF is caused by a variety of genes,some of which have been identified.However,the underlying genetic factors responsible for the majority of MMAF cases are still largely unknown.The glutamine-rich 2(QRICH2)gene plays an essential role in the development of sperm flagella by regulating the expression of essential sperm flagellar biogenesis-associated proteins,and genetic variants of QRICH2 have been identified as the primary cause of MMAF in humans and mice.Here,we recruited a Pakistani consanguineous family to identify the genetic variant causing infertility in patients with MMAF.Whole-exome sequencing and Sanger sequencing were conducted to identify potentially pathogenic variants causing MMAF in infertile patients.Hematoxylin and eosin(HE)staining was performed to analyze sperm morphology.Quantitative polymerase chain reaction,western blot,and immunofluorescence staining analyses were conducted to observe the expression of QRICH2 in spermatozoa.A novel homozygous missense variant(c.4618C>T)in QRICH2 was identified in the affected patients.Morphological analysis of spermatozoa revealed the MMAF phenotype in infertile patients.qPCR revealed a significant reduction in the level of sperm QRICH2 mRNA,and immunofluorescence staining revealed a lack of sperm QRICH2 expression.Additionally,patients harboring a homozygous QRICH2 mutation presented reduced expression of outer dense fiber 2(ODF2)in sperm,whereas sperm expression of A-kinase anchor protein 4(AKAP4)was normal.These findings expand our understanding of the genetic causes of MMAF-associated male infertility and emphasize the importance of genetic counseling.展开更多
Infertility is a global concern,and oligoasthenoteratozoospermia(OAT)is the most severe form of male infertility,characterized by reduced sperm count,decreased motility,and increased abnormal morphology.Multiple morph...Infertility is a global concern,and oligoasthenoteratozoospermia(OAT)is the most severe form of male infertility,characterized by reduced sperm count,decreased motility,and increased abnormal morphology.Multiple morphological abnormalities of the sperm flagella(MMAF)characterize the most severe type of OAT and are usually caused by loss-of-function mutations in the genes essential for vital aspects of sperm biology,including concentration,motility,and morphology.The fibrous sheath interacting protein 2(FSIP2)plays an essential role in sperm flagellar structure and function by regulating such processes as intraflagellar transport and acrosome formation.The present study,employing whole-exome sequencing(WES),identified two FSIP2 mutations in one patient(patient 1),a homozygous missense(c.262C>A,p.P88T)and a homozygous frameshift mutation(c.10948_10951del,p.N3653Nfs*22),as well as a homozygous FSIP2 frameshift mutation(c.15982_15982del,p.I5328Lfs*33)in another patient(patient 2).The results of bioinformatics analysis indicate that the identified missense mutation(c.262C>A)is rare and predicted to have a deleterious effect on FSIP2.Transmission electron microscopy analysis of sperm revealed several abnormalities,including a disorganized mitochondrial sheath,absence of the central pair and some doublets of microtubules,and significant dysplasia of the fibrous sheath.Reverse transcription-polymerase chain reaction(RT-PCR)indicated significantly reduced FSIP2 messenger RNA(mRNA)levels in sperm lysate of the affected individuals.Immunofluorescence staining revealed a complete absence of FSIP2,A-kinase anchor protein 4(AKAP4),sperm-associated antigen 6(SPAG6),intraflagellar transport 20(IFT20)and actin-like 7A(ACTL7A)proteins in the spermatozoa of patients.Thus,the novel FSIP2 variants identified in patient 1 and patient 2 are recognized as pathogenic mutations responsible for MMAF,providing valuable insights for genetic counseling and reproductive decision-making in affected males.展开更多
基金supported by the National Key Research and Development Program of China(No.2021YFC2700202,No.2022YFA0806303,and No.2022YFC2702601)the Global Select Project of the Institute of Health and Medicine,Hefei Comprehensive National Science Center(DJK-LX-2022010)the Joint Fund for New Medicine of USTC(YD9100002034).
文摘Multiple morphological abnormalities of the flagella(MMAF)represent a severe form of sperm defects leading to asthenozoospermia and male infertility.In this study,we identified a novel homozygous splicing mutation(c.871-4 ACA>A)in the adenylate kinase 7(AK7)gene by whole-exome sequencing in infertile individuals.Spermatozoa from affected individuals exhibited typical MMAF characteristics,including coiled,bent,short,absent,and irregular flagella.Transmission electron microscopy analysis showed disorganized axonemal structure and abnormal mitochondrial sheets in sperm flagella.Immunofluorescence staining confirmed the absence of AK7 protein from the patients’spermatozoa,validating the pathogenic nature of the mutation.This study provides direct evidence linking the AK7 gene to MMAF-associated asthenozoospermia in humans,expanding the mutational spectrum of AK7 and enhancing our understanding of the genetic basis of male infertility.
基金supported by the National Key Research and Development Program of China(2021YFC2700202,2022YFA0806303 and 2022YFC2702601)the Global Select Project of the Institute of Health and Medicine,Hefei Comprehensive National Science Center(DJK-LX-2022010)+1 种基金USTC Research Funds of the Double First-Class Initiative(the Joint Fund for New Medicine of USTC)(YD9100002034)the Fundamental Research Funds for the Central Universities(WK9100000004).
文摘Multiple morphological abnormalities of the sperm flagella(MMAF)are characterized by bent,irregular,short,coiled,and absent flagella.MMAF is caused by a variety of genes,some of which have been identified.However,the underlying genetic factors responsible for the majority of MMAF cases are still largely unknown.The glutamine-rich 2(QRICH2)gene plays an essential role in the development of sperm flagella by regulating the expression of essential sperm flagellar biogenesis-associated proteins,and genetic variants of QRICH2 have been identified as the primary cause of MMAF in humans and mice.Here,we recruited a Pakistani consanguineous family to identify the genetic variant causing infertility in patients with MMAF.Whole-exome sequencing and Sanger sequencing were conducted to identify potentially pathogenic variants causing MMAF in infertile patients.Hematoxylin and eosin(HE)staining was performed to analyze sperm morphology.Quantitative polymerase chain reaction,western blot,and immunofluorescence staining analyses were conducted to observe the expression of QRICH2 in spermatozoa.A novel homozygous missense variant(c.4618C>T)in QRICH2 was identified in the affected patients.Morphological analysis of spermatozoa revealed the MMAF phenotype in infertile patients.qPCR revealed a significant reduction in the level of sperm QRICH2 mRNA,and immunofluorescence staining revealed a lack of sperm QRICH2 expression.Additionally,patients harboring a homozygous QRICH2 mutation presented reduced expression of outer dense fiber 2(ODF2)in sperm,whereas sperm expression of A-kinase anchor protein 4(AKAP4)was normal.These findings expand our understanding of the genetic causes of MMAF-associated male infertility and emphasize the importance of genetic counseling.
基金supported by the National Key Research and Developmental Program of China(2022YFC2702601 and 2022YFA0806303)the Global Select Project(DJK-LX-2022010)the Institute of Health and Medicine,Hefei Comprehensive National Science Center,and the Joint Fund for New Medicine of USTC(YD9100002034).
文摘Infertility is a global concern,and oligoasthenoteratozoospermia(OAT)is the most severe form of male infertility,characterized by reduced sperm count,decreased motility,and increased abnormal morphology.Multiple morphological abnormalities of the sperm flagella(MMAF)characterize the most severe type of OAT and are usually caused by loss-of-function mutations in the genes essential for vital aspects of sperm biology,including concentration,motility,and morphology.The fibrous sheath interacting protein 2(FSIP2)plays an essential role in sperm flagellar structure and function by regulating such processes as intraflagellar transport and acrosome formation.The present study,employing whole-exome sequencing(WES),identified two FSIP2 mutations in one patient(patient 1),a homozygous missense(c.262C>A,p.P88T)and a homozygous frameshift mutation(c.10948_10951del,p.N3653Nfs*22),as well as a homozygous FSIP2 frameshift mutation(c.15982_15982del,p.I5328Lfs*33)in another patient(patient 2).The results of bioinformatics analysis indicate that the identified missense mutation(c.262C>A)is rare and predicted to have a deleterious effect on FSIP2.Transmission electron microscopy analysis of sperm revealed several abnormalities,including a disorganized mitochondrial sheath,absence of the central pair and some doublets of microtubules,and significant dysplasia of the fibrous sheath.Reverse transcription-polymerase chain reaction(RT-PCR)indicated significantly reduced FSIP2 messenger RNA(mRNA)levels in sperm lysate of the affected individuals.Immunofluorescence staining revealed a complete absence of FSIP2,A-kinase anchor protein 4(AKAP4),sperm-associated antigen 6(SPAG6),intraflagellar transport 20(IFT20)and actin-like 7A(ACTL7A)proteins in the spermatozoa of patients.Thus,the novel FSIP2 variants identified in patient 1 and patient 2 are recognized as pathogenic mutations responsible for MMAF,providing valuable insights for genetic counseling and reproductive decision-making in affected males.
