The expression characteristics of FAM71D and its association with sperm motility

Abstract What are the features of FAM71D (Family with sequence similarity 71, member D) expression and is there an association between FAM71D expression and sperm motility? FAM71D, a novel protein exclusively expressed in the testis, is located in sperm flagella and is functionally involved in sperm motility. Some testis-specific proteins have been reported as potential diagnostic biomarkers to evaluate the spermatogenesis process and sperm quality. We have identified a novel testis-specific protein, FAM71D, through microarray data analysis, yet little is known about its expression and function. FAM71D mRNA and protein expression was quantified during mouse testis development. Its localization in germ cells was detected by dual-labeled immunostaining in testis sections and sperm smears. The clinical significance was assessed by comparing FAM71D expression in spermatozoa from normozoospermic controls and asthenozoospermic patients. Testes were dissected from C57BL/6 J male mice at postnatal ages of 1, 2, 3, 4, 6, 8 weeks and 6 months, and sperm was collected from cauda epididymides of adult mice by the swim-up method. Human spermatozoa were isolated from 100 human semen samples by density gradient Percoll centrifugation. RT-qPCR and western blot were performed to semi-quantify the expression of FAM71D in mouse testis, and in the ejaculated spermatozoa of normozoospermic controls and asthenozoospermic patients. Immunofluorescence staining was used to detect the localization of FAM71D. Co-immunoprecipitation assay was performed to evaluate the interaction between FAM71D and calmodulin. An antibody blocking assay was employed to assess the role of FAM71D in sperm motility. Our results showed that FAM71D was exclusively expressed in the testis in an age-dependent manner. FAM71D expression exhibited dynamic change in the cytoplasm of spermatids during spermiogenesis and was finally retained in sperm flagella. FAM71D could interact with calmodulin. Use of anti-FAM71D antibody on sperm significantly decreased sperm motility. Expression level of FAM71D was markedly reduced in the ejaculated spermataozoa of asthenozoospermic patients (P