Characterization of Cherax quadricarinatus prohibitin and its potential role in spermatogenesis

Prohibitin (PHB) proteins have diverse functions, such as cellular signaling, transcriptional control and mitochondrial biogenesis. In this study, we characterized PHB gene and its protein expression in Cherax quadricarinatus. PHB cDNA comprises 1472 nucleotides with an open reading frame of 828 bp, which encodes 275 amino acid residues. The highest transcript levels were found during the spermatogonial developmental phase, with the lowest levels detected during the resting phase in the reproductive cycle. Western blot analysis revealed that PHB is an approximately 30 kDa protein, and occurs in a number of unexpected isoforms, ranging from 30 kDa to greater than 180 kDa in the testes of different developmental phases, which may be the ubiquitinated substrates. The strongest immunolabeling signal was found in spermatogonia, with lower levels of staining in secondary spermatocytes, and weak or absent expression in mature sperm. Immunogold electron microscopy results confirmed the localization of PHB in the inner mitochondrial membranes. The results showed that PHB is a substrate protein for spermatogenesis, with a potential reproductive function involving sperm ubiquitination in invertebrates.