Immunogold electron microscopic localization of antigenic sites in the outer dense fiber region of rat sperm tail obtained by using antisera to two Sertoli cell secretory proteins

Abstract

Polyclonal antisera raised against polypeptide components of two rat Sertoli cell secretory proteins, designated protein S70 and S45-S35 heterodimeric protein on the basis of cell origin and estimated molecular weight, were used to identify antigenic sites in 1) rat testis, 2) cultured Sertoli cells, 3) developing spermatids (collected from spermatogenic stage-specific seminiferous tubular segments), and 4) epididymal sperm. Indirect immunofluorescence, immunoperoxidase, and immunogold electron microscopy (single and double labeling) were used. Immunocytochemical techniques have detected antigenic sites in 1) the cytoplasm of Sertoli cells in the intact seminiferous tubule and in culture in the form of a punctuate, granular-like pattern, and 2) the acrosome (but not the Golgi region) and tail of developing spermatids and sperm. In developing spermatids, the principal piece of the tail displays a characteristic apical-to-distal immunoreactive banding pattern that correlates both temporally and spatially with the reported multistep assembly of outer dense fibers along the axoneme. The immunoreactivity of the acrosome, connecting piece, and outer dense fibers of the sperm tail was confirmed by immunogold electron microscopy. A precise identification of the component(s) of the outer dense fiber region responsible for the antigenic homology with Sertoli cell secretory proteins is under investigation.

Rat sertoli and spermatogenic cells express a similar gene, and its product is antigenically related to an outer dense fiber‐associated protein

“…Supporting our previous observations (Kierszenbaum et al, 1991), we demonstrate that a protein associated with the extracted outer dense fiber fraction, not the outer dense fibers themselves, is responsible for the antigenic homology. The following results support our interpretation: 1) immunogold electron microscopic experiments using whole-mounted outer dense fibers showed strong immunoreactivity on the surface of the extracted outer dense fibers.…”

“…The presence of specific mRNA transcripts and encoded protein in primary spermatocytes and spermatids that are antigenically related to Sertoli cell heterodimeric protein suggest a function of this gene during spermatogenesis. Previous work has shown that specific antisera to Sertoli cell heterodimeric protein detected antigenically related proteins in the acrosome and outer dense fiber region of the tail of developing spermatids and sperm (Abdullah et al, 1988;Kierszenbaum et al, 1988Kierszenbaum et al, , 1989Kierszenbaum et al, , 1991. It was unclear from these observations whether antigenically homologous proteins 1) derived from Sertoli cells and then translocated to spermatogenic cells or 2) were synthesized de novo by spermatogenic cells.…”

“…Immunoblotting and immunocytochemical experiments have shown that antisera to the 73 kD precursor of Sertoli cell heterodimeric protein and to the 45 kD and 35 kD components of the heterodimer cross react with a component of the outer dense fibers of rat sperm tail (Abdullah et al, 1988;Kierszenbaum et al, 1988Kierszenbaum et al, , 1989Kierszenbaum et al, ,1991. Polyclonal antisera generated against several protein components of extracted outer dense fibers immunoprecipitated Sertoli cell 73,45, and 35 kD proteins (Kierszenbaum et al, 1989).…”

“…The diversity of names and functions ascribed to products expressed by this widespread gene suggests important physiological and structural roles. ' We have reported that components of the acrosome and the outer dense fiber region of the tail of developing spermatids and mature sperm show antigenic homology to the heterodimeric protein and its precursor (Abdullah et al, 1988;Kierszenbaum et al, 1988Kierszenbaum et al, , 1989Kierszenbaum et al, , 1991. Furthermore, antibodies to polypeptides extracted from the outer dense fiber fraction of sperm tail cross react with the heterodimeric protein and its precursor (Kierszenbaum et al, 1989).…”

Developmental expression of the S35‐S45/SGP‐2/TRPM‐2 gene in rat testis and epididymis

“…The protein is extensively modified after translation, and there is evidence that these posttranslational modifications may vary quite significantly from tissue to tissue (Buttyan et al, 1989;Cheng et al, 1990). Antibodies to the subunit components of SGP-2iS45-35 and to its precursor, S70, recognize antigenic sites in the acrosome and the outer dense fiber region of the tail of developing spermatids and sperm Kierszenbaum et al, 1989Kierszenbaum et al, ,1991. To gain insight into the function of TRPM-2, we have characterized the expression of the gene in rat testis and epididymis during development.…”