Identification of a nuclear localization signal in activin/inhibin βA subunit; intranuclear βA in rat spermatogenic cells

Activin is a dimeric glycoprotein hormone that was initially characterized by its ability to stimulate pituitary FSH secretion and was subsequently recognized as a growth factor with diverse biological functions in a large variety of tissues. In the testis, activin has been implicated in the auto/paracrine regulation of spermatogenesis through its cognate cell membrane receptors on Sertoli and germ cells. In this study we provide evidence for intranuclear activin/inhibin beta(A) subunit and show its distribution in the rat seminiferous epithelium. We have shown by transient expression in HeLa cells of beta-galactosidase fusion proteins that the beta(A) subunit precursor contains a functional nuclear localization signal within the lysine-rich sequence corresponding to amino acids 231-244. In all stages of the rat seminiferous epithelial cycle, an intense immunohistochemical staining of nuclear beta(A) was demonstrated in intermediate or type B spermatogonia or primary spermatocytes in their initial stages of the first meiotic prophase, as well as in pachytene spermatocytes and elongating spermatids primarily in stages IX-XII. In some pachytene spermatocytes, the pattern of beta(A) immunoreactivity was consistent with the characteristic distribution of pachytene chromosomes. In the nuclei of round spermatids, beta(A) immunoreactivity was less intense, and in late spermatids it was localized in the residual cytoplasm, suggesting disposal of beta(A) before spermatozoal maturation. Immunoblot analysis of a protein extract from isolated testicular nuclei revealed a nuclear beta(A) species with a molecular mass of approximately 24 kDa, which is more than 1.5 times that of the mature activin beta(A) subunit present in activin dimers. These results suggest that activin/inhibin beta(A) may elicit its biological functions through two parallel signal transduction pathways, one involving the dimeric molecule and cell surface receptors and the other an alternately processed beta(A) sequence acting directly within the nucleus. According to our immunohistochemical data, beta(A) may play a significant role in the regulation of nuclear functions during meiosis and spermiogenesis.