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Caveolae are 50-100 nm flask-shaped invaginations of the plasma membrane found in most cell types. Caveolin-1 is the principal protein component of caveolae membranes in nonmuscle cells. The recent development of Cav-1-deficient mice has allowed investigators to study the in vivo functional role of caveolae in the context of a whole animal model, as these mice lack morphologically detectable caveolae membrane domains. Surprisingly, Cav-1 null mice are both viable and fertile. However, it remains unknown whether loss of caveolin-1 significantly affects the overall life span of these animals. To quantitatively determine whether loss of Cav-1 gene expression confers any survival disadvantages with increasing age, we generated a large cohort of mice (n = 180), consisting of Cav-1 wild-type (+/+) (n = 53), Cav-1 heterozygous (+/-) (n = 70), and Cav-1 knockout (-/-) (n = 57) animals, and monitored their long-term survival over a 2 year period. Here, we show that Cav-1 null (-/-) mice exhibit an approximately 50% reduction in life span, with major declines in viability occurring between 27 and 65 weeks of age. However, Cav-1 heterozygous (+/-) mice did not show any changes in long-term survival, indicating that loss of both Cav-1 alleles is required to mediate a reduction in life span. Mechanistically, these dramatic reductions in life span appear to be secondary to a combination of pulmonary fibrosis, pulmonary hypertension, and cardiac hypertrophy in Cav-1 null mice. Taken together, our results provide the first demonstration that loss of Cav-1 gene expression and caveolae organelles dramatically affects the long-term survival of an organism. In addition, aged Cav-1 null mice may provide a new animal model to study the pathogenesis and treatment of progressive hypertrophic cardiomyopathy and sudden cardiac death syndrome.

Caveolae are vesicular invaginations of the plasma membrane. Their formation is strictly dependent on the expression of the caveolin coat proteins. During transit to the plasma membrane, approximately 15 monomers of caveolin-1 assemble into a multivalent homo-oligomer. Caveolae are most likely generated through the subsequent interaction of these caveolin homo-oligomers with one another, with sphingolipids, and with cholesterol. Membrane association of caveolin-1 is critical to this process and is facilitated by an atypical N-terminal membrane attachment domain (residues 82-101), termed N-MAD. To better understand the membrane attachment function of N-MAD, we performed a detailed mutational analysis of the 20 amino acid N-MAD peptide sequence fused to the C-terminus of the soluble reporter green fluorescent protein (GFP). Removal of the distal six residues (KYWFYR) within N-MAD prevents membrane attachment in cells as assessed by hypotonic lysis, detergent solubility, carbonate extraction, and fluorescence microscopy. These six residues (KYWFYR) are sufficient to confer membrane attachment to GFP, an otherwise soluble protein. Both the central aromatic and flanking basic residues in this sequence are required for membrane attachment, as the sequence YWFY does not confer membrane affinity to GFP. Although the KYWFYR sequence within N-MAD facilitates membrane association, we show that the entire N-MAD sequence is required for targeting to lipid rafts/caveolae.