Targeting of the domain-specific integral membrane protein PM52 to the periacrosomal plasma membrane during guinea pig spermiogenesis.

Abstract

The sperm plasma membrane is segregated into functionally, biochemically, and structurally distinct domains yet the protein sorting pathways and assembly mechanisms that assemble these domains during spermiogenesis are incompletely understood. We previously characterized two structurally related size-variant, integral membrane proteins of 52 kDa (PM52) and 35 kDa localized to the periacrosomal plasma membrane of guinea pig cauda epididymal spermatozoa (Westbrook-Case et al., 1994). In this study we used light and electron microscopic immunocytochemistry to define the expression pattern and sorting pathway that establishes the domain-specific distribution of PM52 during spermiogenesis. The PM52 is first expressed in acrosome-phase spermatids and it localizes exclusively to the cytoplasmic lobe. Immunoelectron microscopy revealed that both cytoplasmic vesicles and the plasma membrane of the cytoplasmic lobe labeled with anti-PM52. During early stages of expression, PM52 appeared to be absent from the head region, but significant PM52 accumulation over the spermatid head was noted in late acrosomal phase spermatids. Throughout spermiogenesis PM52 extended posteriorly to the annulus, which represents a barrier preventing PM52 diffusion into the posterior tail. Following the migration of the annulus to the midpiece-principal piece junction, PM52 began to disappear from the flagellar region and at the completion of spermiogenesis most of the PM52 was restricted to the acrosomal segment. Spermatids and epididymal sperm PM52 exhibited identical sizes by SDS-PAGE and immunoblotting, indicating that they are not proteolytically modified during epididymal maturation. The PM52 antibodies were also used to screen a guinea pig testis cDNA library, and sequence determination of full-length PM52 clones demonstrated identity of a sperm membrane protein recently termed "sperad" (Quill and Garbers, 1996). Membrane barriers and potential mechanisms establishing the domain-specific residence of PM52 are discussed.