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First published online February 22, 2006
doi: 10.1242/10.1242/jcs.02853

Journal of Cell Science 119, 787-796 (2006)
Published by The Company of Biologists 2006
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Biogenesis of caveolae: a structural model for caveolin-induced domain formation

Robert G. Parton1,2,*, Michael Hanzal-Bayer1 and John F. Hancock1

1 Institute for Molecular Bioscience, University of Queensland, Queensland 4072, Australia
2 Centre for Microscopy and Microanalysis, University of Queensland, Queensland 4072, Australia


Figure 1
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  		Fig. 1. Caveolae and caveolin structure. Main panel: caveolae in a differentiated 3T3-L1 adipocyte labelled with an electron-dense marker to delineate the cell surface. Note the uniform size and shape of caveolae (some are indicated by arrowheads) and the numerous caveolae around larger surface-connected vacuoles (asterisks). Lower left: caveolae (arrowheads) in a primary fibroblast shown in a conventional plastic section. Top right: a similar image at higher magnification. Lower right: sheets of plasma membrane from adipocytes labelled for caveolin-1 followed by 10 nm protein-A-gold. Note the unlabeled clathrin-coated structure (double arrowheads). The schematic shows the main features of caveolin association with the plasma membrane; positively charged (+) and aromatic (·) amino acids in the scaffolding domain interact strongly with the membrane; schematic modified from Arbuzova et al. (Arbuzova et al., 2000Go). Cholesterol may be enriched in the scaffolding-domain-associated portion of the membrane. Bars, 100 nm.

 

Figure 2
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  		Fig. 2. The caveolae `coat'. The images show the spiked caveolar coat (left; red dots indicate spikes) as compared with the classical clathrin coat (right) in sections of baby hamster kidney (BHK) cells. The ultrathin plastic sections (<30 nm thickness) allow detection of the coat, which is normally difficult to visualise by conventional EM techniques (compare with Fig. 1). Bars, 100 nm.

 

Figure 3
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  		Fig. 3. Sequence features and disease-associated amino acid changes in caveolins-1 (Cav1) and -3 (Cav3). Some key amino acids and features of caveolin-1 and caveolin-3 are shown in relation to defined domains of caveolin-1 and caveolin-3. Numbers above the lines indicate amino acid number in mammalian caveolins. Palmitoylation sites (Palm) in caveolin-1 are indicated in green (potential sites in caveolin-3 are not shown), phosphorylation sites (P) are shown in red, and the starting methionine (M33) of caveolin-1ß is shown in black. Caveolin-3 residues are numbered assuming a single methionine residue at the N-terminus. Disease-associated amino acid substitutions in caveolin-1 and caveolin-3 are shown in blue. Abbreviations: RMD, rippling muscle disease; DM, distal myopathy; FHCK, familial hyperCKaemia; SHCK, spontaneous hyperCKaemia; MYO, myopathy; LGMD1C, limb girdle muscular dystrophy 1C; BC, breast cancer.

 

Figure 4
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  		Fig. 4. Topology model of caveolin-1. Helical and buried regions have been predicted from primary structure. The model assumes that charged groups (green) have access to the membrane surface and tryptophan residues (W) are hydrogen bonded. Note that tryptophan residues stay within one helical turn of either the cytosolic (W85, W98, W128) or the exterior (W115) membrane interface. Aromatic residues tend to stay together (black numbers indicate their position in rotational degrees relative to the first aromatic residue of each segment). Segment 80-95 qualifies as an in-plane amphipathic membrane anchor and might compensate for the effect of segment 95-110 being shorter than segment 110-130. Aromatic residues are depicted in yellow, known surface accessible residues in red, and the palmitoyl moieties are to scale. (Numbers correspond to murine caveolin-1.)

 

Figure 5
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  		Fig. 5. Model of volume contributions from caveolin and cholesterol in caveolae. The model estimates the increase in volume of the hydrophobic core of the cytoplasmic leaflet (Vc) of the plasma membrane compared with the hydrophobic core of the outer leaflet (Vo) that would be required to cause the curvature characteristic of a caveola. The diagram shows the ellipsoid shape of a fast-frozen adipocyte caveola for which we obtained values of a=42 nm, b=33 nm, c=21 nm, h=29 nm. Vc and Vo were calculated by sequentially subtracting the volumes of a set of three concentric ellipsoids obtained by reducing the dimensions of a, b and c by t and 2t, where t is the thickness of the hydrophobic core of a single leaflet (t=1.5 nm). The volume of a solid ellipsoid of the configuration shown is given by V={pi}/3 (2ab2+2hb2+hc2). We assume that only the scaffolding domain helix {alpha}2a contributes to expansion of Vc because the two membrane-spanning helices ({alpha}2b and helix {alpha}3) contribute equal volumes to Vc and Vo. We further assume that there are 145 caveolin molecules per caveola (Ncav) and that the volume of {alpha}2a, V{alpha}2a=2.2 nm3. We propose that the additional volume required to expand the cytoplasmic leaflet (Vc-Vo) is contributed by the ability of the scaffolding domain to penetrate the hydrophobic core of the cytoplasmic leaflet and sequester cholesterol in an asymmetric fashion across the bilayer. Nchol is the number of cholesterol molecules sequestered by caveolin, and we assume that the volume of a cholesterol molecule is 0.612 nm3. In this model, the volume contributed by cholesterol, Vchol, would correspond to a molecular ratio of cholesterol to caveolin of 13:1.

 

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© The Company of Biologists Ltd 2006