Voltage-dependent Anion-selective Channels VDAC2 and VDAC3 Are Abundant Proteins in Bovine Outer Dense Fibers, a Cytoskeletal Component of the Sperm Flagellum

Abstract

Outer dense fibers (ODF) are specific subcellular components of the sperm flagellum. The functions of ODF have not yet been clearly elucidated. We have investigated the protein composition of purified ODF from bovine spermatozoa and found that one of the most abundant proteins is a 30 -32-kDa polypeptide. This protein was analyzed by sequencing peptides derived following limited proteolysis. Peptide sequences were found to match VDAC2 and VDAC3. VDACs (voltage-dependent, anion-selective channels) or eukaryotic porins are a group of proteins first identified in the mitochondrial outer membrane that are able to form hydrophilic pore structures in membranes. In mammals, three VDAC isoforms (VDAC1, -2, -3) have been identified by cDNA cloning and sequencing. Antibodies against synthetic peptides specific for the three mammal VDAC isoforms were generated in rabbits. Their specificity was demonstrated by immunoblotting using recombinant VDAC1, -2, and -3. In protein extracts of bovine spermatozoa, VDAC1, -2, and -3 were detected by specific antibodies, while only VDAC2 and -3 were found as solubilized proteins derived from purified bovine ODFs. Immunofluorescence microscopy of spermatozoa revealed that anti-VDAC2 and anti-VDAC3 antibodies clearly bound to the sperm flagellum, in particular to the ODF. Transmission electron immunomicroscopy supported the finding that VDAC2 protein is abundant in the ODF. Since the ODF does not have any known membranous structure, it is tempting to speculate that VDAC2 and VDAC3 might have an alternative structural organization and different functions in ODF than in mitochondria.

Novelty of the Pyruvate Metabolic Enzyme Dihydrolipoamide Dehydrogenasein Spermatozoa

“…Phospholipid hydroperoxide glutathione peroxidase (PHGPx) is a cytosolic protein in somatic cells, which is found to be associated with the mitochondria in the mature spermatozoa (50). On the other hand, Voltage-dependent anion-selective channels (VDAC2 and VDAC3), which are mitochondrial porins in somatic cells, are found to be associated with the cytoskeletal elements (outer dense fibers) in the bovine sperm flagella (60). It is obvious that such dramatic changes in localization probably imply fulfillment of specific functions.…”

Swapping of the N‐terminus of VDAC1 with VDAC3 restores full activity of the channel and confers anti‐aging features to the cell

“…mice were generated. VDAC1 deficient mice are viable and show that mitochondrial functions are slightly affected; VDAC2 gene deletion is lethal [14]; the spermatozoa are rich in VDAC3 [15]: mice lacking VDAC3 are healthy, but males are infertile [16].…”

Analysis and difference of voltage-dependent anion channel mRNA in ejaculated spermatozoa from normozoospermic fertile donors and infertile patients with idiopathic asthenozoospermia

“…Few recent studies have reported that VDAC is present in mammalian spermatozoa and plays putative roles in spermatogenesis, sperm maturation and fertilization [15][16][17][18][19]. However the respective expression, localization and function of three VDAC subtypes remain uncertain.…”