Cullin-RING E3 ubiquitin ligase (CRL)-4 is a member of the large CRL family in eukaryotes. It plays important roles in a wide range of cellular processes, organismal development, and physiological and pathological conditions. DDB1- and CUL4-associated factor 8 (DCAF8) is a WD40 repeat-containing protein, which serves as a substrate receptor for CRL4. The physiological role of DCAF8 is unknown. In this study, we constructed knockout mice. Homozygous mice were viable with no noticeable abnormalities. However, the fertility of deficient male mice was markedly impaired, consistent with the high expression of DCAF8 in adult mouse testis. Sperm movement characteristics, including progressive motility, path velocity, progressive velocity, and track speed, were significantly lower in knockout mice than in wild-type (WT) mice. However, the total motility was similar between WT and knockout sperm. More than 40% of spermatids in knockout mice showed pronounced morphological abnormalities with typical bent head malformation. The acrosome and nucleus of knockout sperm looked similar to those of WT sperm. tests showed that the fertilization rate of knockout mice was significantly reduced. The results demonstrated that DCAF8 plays a critical role in spermatogenesis, and DCAF8 is a key component of CRL4 function in the reproductive system.