cDNA sequence analysis, gene expression and protein localisation of the inhibin alpha-subunit of Australian brushtail possum (Trichosurus vulpecula)

Abstract

An inhibin alpha-subunit cDNA sequence from the Australian brushtail possum (Trichosurus vulpecula) has been identified and analysed. The cDNA includes an open reading frame encoding a predicted precursor protein of 361 amino acids. The predicted protein sequence includes four possible proteolytic cleavage sites, 12 evolutionarily conserved cysteine residues and three potential N-linked glycosylation sites. The mature alpha-subunit is the carboxyl terminal fragment (alphaC) consisting of 131 amino acids. The full-length precursor protein shows a mean identity with eutherian homologues of 69.8%. The homology is not evenly distributed, with the putative alphaC fragment showing the highest level (79.7%). Using Northern hybridisation, an alpha-subunit transcript of approximately 1.6 kb was detected in adult possum ovary. Using in situ hybridisation and immunocytochemistry, inhibin alpha-subunit was localised exclusively to the granulosa cell layers of follicles. Hybridisation and immunostaining for the inhibin alpha-subunit were first observed in granulosa cells of primary follicles and the expression continued throughout all stages of follicular growth. Inhibin alpha-subunit mRNA and protein were also detected in cells of the corpus luteum. In summary, results indicate considerable conservation of the structure and possible function of the inhibin alpha-subunit protein since the divergence of the marsupial and eutherian mammalian lineages. The expression data suggest that, in the adult possum, inhibin may have a role in ovarian follicular growth from the primary stage of development.