A loss-of-function variant in ZCWPW1 causes human male infertility with sperm head defect and high DNA fragmentation

Abstract

Abstract Background In about one-third of cases, the genetic causes of asthenozoospermia are unknown. The more causative genes related to human male infertility should be further explored. The essential role of ZCWPW1 in mouse male fertility has been established and the role of ZCWPW1 in human reproduction need further investigation to verify. Methods Whole-exome sequencing was conducted to identify causative genes in the infertile man. The phenotype of the asthenozoospermia was determined by Papanicolaou staining, immunofluorescence staining and electron microscope. In-vitro experiments, western blot and in-silicon analysis were applied to assess the pathogenicity of the identified variant. Additionally, we examined the influence of the variant on the DNA fragmentation and DNA repair capability by Sperm Chromatin Dispersion and Neutral Comet Assay. Results The spermatozoa of the proband exhibited low quantity, head defects in semen examination, papanicolaou staining and electron microscope. Whole-exome sequencing and sanger sequencing found the proband carried a homozygous ZCWPW1 variant (c.1064C > T, p. P355L). Immunofluorescence analysis showed that a significant decrease of ZCWPW1 expression in proband’s sperm. The obvious declined expression of ZCWPW1 with the mutation was validated in HEK293T. Furthermore, Sperm Chromatin Dispersion assay revealed the proband’s spermatozoa had high DNA fragmentation. After treated by hydroxyurea, MUT-ZCWPW1 transfected cells and empty vector transfected cells had higher level of γ-H2AX and tail DNA than WT-ZCWPW1 transfected cells. Conclusions It is the first report that a novel homozygous missense mutation in ZCWPW1 caused human male infertility with sperm head defect and high DNA fragmentation. This finding enriches the gene variant spectrum and etiology of asthenozoospermia.