Study On The Mechanism Of Acephalic Spermatozoa Syndrome Caused By A Novel Mutation In PMFBP1

Infertility is an increasingly serious health problem worldwide.About 15% of couples of childbearing age suffer from infertility,and about 50% of them are caused by male factors.Therefore,the problem of male infertility has attracted more and more social attention,and teratozoospermia is one of the main causes of male infertility.There are many types of teratozoospermia,among which acephalic spermatozoa syndrome is a serious and rare teratozoospermia.The main clinical feature of acephalic spermatozoa syndrome is the presence of a large number of headless sperm tails in semen,accompanied by a small number of tailless sperm heads and sperm with abnormal head-to-tail connections.This abnormality of headless sperm is also called decapitated sperm or pin-like sperm.The pathogenic genes of acephalic spermatozoa syndrome that have been reported include SUN5,PMFBP1,BRDT,HOOK1,TSGA10,DNAH6,CEP112,but there are still some cases that cannot be explained by reported gene mutations.Therefore,new pathogenic genes have been discovered or novel mutation is of great significance for he diagnosis and treatment of acephalic spermatozoa syndrome,and it also helps to clarify the pathogenic mechanism of acephalic spermatozoa syndrome.This paper recruited a patient with acephalic spermatozoa syndrome from a consanguineous family,except for infertility,other clinical examination indicators were within the range.First,the patient’s peripheral blood was drawn to extract DNA to screen for mutations in the main pathogenic genes SUN5 and PMFBP1 and a novel homozygous missense mutation in exon 4 of PMFBP1 was found,c.301A> C;(p.T101P).Secondly,the pathogenicity of the mutation was predicted by prediction softwares such as SIFT,Polyphen-2,Mutation Taster;and the mutation was not detected in databases such as db SNP153,gnom AD and Ex AC.Finally,the expression and localization of PMFBP1 in the sperm of patients were detected by Western blot and immunofluorescence.At the same time,a mutant of PMFBP1 was constructed,and the effect of mutation on the expression level and localization of PMFBP1 protein was further analyzed through cell transfection experiments.The experimental results showed that the expression of the PMFBP1 mutant protein in the sperm of the acephalic spermatozoa syndrome patient was significantly reduced compared with the normal control.In vitro cell experiment also showed that the expression level of PMFBP1 mutant protein in sperm was significantly reduced.It is speculated that the homozygous missense mutation c.301A>C(p.T101P)of PMFBP1 may be the cause of the patient with acephalic spermatozoa syndrome.In addition,we collected 3 pmfbp1 knockout and 3 wild-type adult mouse testis samples for quantitative proteomics analysis.Proteomics identified a total of 7,315 proteins,of which 6,511 proteins contained quantitative information,and 125 of them were down-regulated Protein,13 up-regulated proteins.We conducted a systematic bioinformatics analysis(protein function annotation)of all identified proteins,and performed functional classification,functional enrichment,and cluster analysis based on functional enrichment of all differentially expressed proteins to obtain the main influence of the gene The sperm capacitation,the integrity and stability of the sperm structure are improved.The novel mutation of PMFBP1 and the differential protein profile of proteomics provide the basis for genetic counseling and clinical diagnosis and treatment of patients with acephalic spermatozoa syndrome.