Voltage-dependent anion-selective channels VDAC2 and VDAC3 are abundant proteins in bovine outer dense fibers, a cytoskeletal component of the sperm flagellum.
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.
Hinsch
Klaus-Dieter
KD
Center of Dermatology and Andrology, Justus Liebig University, 35392 Giessen, Germany. Klaus-Dieter.Hinsch@derma.med.uni-giessen.de
De Pinto
Vito
V
Aires
Viviana A
VA
Schneider
Xenia
X
Messina
Angela
A
Hinsch
Elvira
E
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D003599:Cytoskeleton / Q000378:metabolism*; D018076:DNA, Complementary / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D005407:Flagella / Q000378:metabolism*; D006801:Humans; D015151:Immunoblotting; D007425:Intracellular Membranes / Q000378:metabolism; D008297:Male; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000378:metabolism; D033681:Mitochondrial Membrane Transport Proteins; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D016133:Polymerase Chain Reaction; D018272:Porins / Q000737:chemistry*; D011485:Protein Binding; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013094:Spermatozoa / Q000378:metabolism*; D050995:Voltage-Dependent Anion Channel 1; D050996:Voltage-Dependent Anion Channel 2; D050994:Voltage-Dependent Anion Channels
D018076:DNA, Complementary; D033681:Mitochondrial Membrane Transport Proteins; D010455:Peptides; D018272:Porins; D020033:Protein Isoforms; D011994:Recombinant Proteins; C494762:VDAC1 protein, human; C494765:VDAC2 protein, human; C494768:VDAC3 protein, human; D050996:Voltage-Dependent Anion Channel 2; D050994:Voltage-Dependent Anion Channels; D050995:Voltage-Dependent Anion Channel 1