Prohibitin Protein Expression During Spermatogenesis in the Large Yellow Croaker, Larimichthys crocea

Mitochondria are important for animals’ fertility, and their morphologies and functions during spermatogenesis are under investigation. However, the molecular mechanism that regulates the mitochondrial dynamic during spermatogenesis is still unknown. In this study, the cytological features of spermatogenesis were investigated in Larimichthys crocea. In addition, the structure and function of prohibitin (PHB), which is associated with mitochondrial structure and dynamic, was also investigated. The full-length cDNA and protein ( Lc -PHB) from the L. crocea phb gene ( Lc-phb ) contained 1625 base pairs and 271 amino acids, respectively. Lc -PHB had a conserved primary structure that resulted in a transmembrane, SPFH (the analogous region of proteins stomatins, prohibitins, flotillins and HflK/C), and coiled-coil domains. It was detected at high levels in the muscle, liver, and heart, and at intermediate levels in the testis, gill, and brain. Lc-phb mRNA expression was detected in spermatogenic cells by fluorescence in situ hybridization. An immunofluorescence assay revealed that PHB protein was localized in the mitochondria during spermatogenesis. Specifically, PHB expression was detected in the perinuclear cytoplasm of spermatogonia, spermatocytes, and spermatids in the early developmental stage, and mainly localized on one side of the nuclei in the cytoplasm of spermatids in a middle developmental stage, and finally on the sperm midpiece. Western blotting showed that PHB was located in the extracted mitochondria protein fraction but not in the cytoplasm protein fraction of testes. Conclusively, these results indicated that PHB was expressed in the mitochondria during spermatogenesis. In addition, the study explained the mitochondrial dynamic during fish spermatogenesis and proposed a possible relationship among PHB, spermatogenesis, and male fertility.