Expression of the human antigen SPAG2 in the testis and localization to the outer dense fibers in spermatozoa

Antisperm antibodies (ASAs) have been implicated in some instances of infertility. To characterize sperm antigens relevant to immunologic and immunocontraceptive development, SPAG2 (sperm-associated antigen 2) was identified by screening a human testis cDNA library with human sera positive for ASAs. Subsequently, two isoforms, SPAG2-1 and SPAG2-2, were identified in testis and placenta libraries, respectively. In the current study, Southern analysis of human genomic DNA with a probe common to the two SPAG2 isoforms indicated a single SPAG2 gene; therefore, alternative splicing is a likely mechanism for production of variant mRNAs. In situ hybridization of human testis sections demonstrated the expression of SPAG2 in primary spermatocytes, with decreased or arrested expression in postmeiotic cells. Immunofluorescence of Triton X-100-extracted spermatozoa with an anti-SPAG2 peptide antiserum indicate that SPAG2 is an intracellular component of the sperm flagellum. Electron microscopy refined this localization to the outer dense fibers (ODFs), structural filaments associated with the mammalian sperm axoneme. The ODFs have been reported to be composed of keratin-like intermediate filament proteins. However, SPAG2 does not exhibit the molecular characteristics of such proteins, nor does SPAG2 demonstrate sequence homology with previously characterized ODF proteins. Therefore, SPAG2 represents a novel protein of human sperm ODFs. Characterization of SPAG2 will further our understanding of ODF function in normal sperm motility and of flagellar abnormalities that lead to male infertility.