A novel protein, sperm head and tail associated protein (SHTAP), interacts with cysteine-rich secretory protein 2 (CRISP2) during spermatogenesis in the mouse

Abstract

CRISP2 (cysteine-rich secretory protein 2) is a sperm acrosome and tail protein with the ability to regulate Ca2+ flow through ryanodine receptors. Based on these properties, CRISP2 has a potential role in fertilization through the regulation of ion signalling in the acrosome reaction and sperm motility. The purpose of the present study was to determine the expression, subcellular localization and the role in spermatogenesis of a novel CRISP2-binding partner, which we have designated SHTAP (sperm head and tail associated protein).Using yeast two-hybrid screens of an adult testis expression library, we identified SHTAP as a novel mouse CRISP2-binding partner. Sequence analysis of all Shtap cDNA clones revealed that the mouse Shtap gene is embedded within a gene encoding the unrelated protein NSUN4 (NOL1/NOP2/Sun domain family member 4). Five orthologues of the Shtap gene have been annotated in public databases. SHTAP and its orthologues showed no significant sequence similarity to any known protein or functional motifs, including NSUN4. Using an SHTAP antiserum, multiple SHTAP isoforms (approximately 20-87 kDa) were detected in the testis, sperm, and various somatic tissues. Interestingly, only the approximately 26 kDa isoform of SHTAP was able to interact with CRISP2. Furthermore, yeast two-hybrid assays showed that both the CAP (CRISP/antigen 5/pathogenesis related-1) and CRISP domains of CRISP2 were required for maximal binding to SHTAP. SHTAP protein was localized to the peri-acrosomal region of round spermatids, and the head and tail of the elongated spermatids and sperm tail where it co-localized with CRISP2. During sperm capacitation, SHTAP and the SHTAP-CRISP2 complex appeared to be redistributed within the head.The present study is the first report of the identification, annotation and expression analysis of the mouse Shtap gene. The redistribution observed during sperm capacitation raises the possibility that SHTAP and the SHTAP-CRISP2 complex play a role in the attainment of sperm functional competence.