VIP21-CAVEOLIN, A MEMBRANE-PROTEIN CONSTITUENT OF THE CAVEOLAR COAT, OLIGOMERIZES IN-VIVO AND IN-VITRO

VIP21-caveolin is a membrane protein, proposed to be a component of th e striated coat covering the cytoplasmic surface of caveolae. To inves tigate the biochemical composition of the caveolar coat, we used our p revious observation that VIP21-caveolin is present in large complexes and insoluble in the detergents CHAPS or Triton X-114. The mild treatm ent of these insoluble structures with sodium dodecyl sulfate leads to the detection of high molecular mass complexes of approximately 200, 400, and 600 kDa. The 40O-kDa complex purified to homogeneity from dog lung is shown to consist exclusive of the two isoforms of VIP21-caveo lin. Pulse-chase experiments indicate that the oligomers form early af ter the protein is synthesized in the endoplasmic reticulum (ER). VIP2 1-caveolin does indeed insert into the ER membrane through the classic al translocation machinery. Its hydrophobic domain adopts an unusual l oop configuration exposing the N- and C-flanking regions to the cytopl asm. Similar high molecular mass complexes can be produced from the in vitro-synthesized VIP21-caveolin. The complex formation occurs only i f VIP21-caveolin isoforms are properly inserted into the membrane; for mation is cytosol-dependent and does not involve a vesicle fusion step We propose that high molecular mass oligomers of VIP21-caveolin repre sent the basic units forming the caveolar coat. They are formed in the ER and later, between the ER and the plasma membrane, these oligomers could associate into larger detergent-insoluble structures.