Regulation of cAMP-mediated signal transduction via interaction of caveolins with the catalytic subunit of protein kinase A

cAMP-dependent processes are essential for cell growth, differentiation, an d homeostasis. The classic components of this system include the serpentine receptors, heterotrimeric G-proteins, adenylyl cyclase, protein kinase A ( PKA), and numerous downstream target substrates. Evidence is accumulating t hat some members of this cascade are concentrated within membrane microdoma ins, termed caveolae and caveolae-related domains. In addition, the caveoli n-1 protein has been shown to interact with some of these components, and t his interaction inhibits their enzymatic activity, However, the functional effects of caveolins on cAMP-mediated signaling at the most pivotal step, P EA activation, remain unknown. Here, we show that caveolin-1 can dramatical ly inhibit cAMP-dependent signaling in vivo. We provide evidence for a dire ct interaction between caveolin-1 and the catalytic subunit of PKA both in vitro and in vivo. Caveolin-1 binding appears to be mediated both by the ca veolin scaffolding domain (residues 82-101) and a portion of the C-terminal domain (residues 135-156), Further functional analysis indicates that cave olin-based peptides derived from these binding regions can inhibit the cata lytic activity of purified PKA in vitro. Mutational analysis of the caveoli n scaffolding domain reveals that a series of aromatic residues within the caveolin scaffolding domain are critical for mediating inhibition of PKA, I n addition, co-expression of caveolin-1 and PKA in cultured cells results i n their co-localization as seen by immunofluorescence microscopy. In cells co-expressing caveolin-1 and PKA, PKA assumed a punctate distribution that coincided with the distribution of caveolin-1. In contrast, in cells expres sing PKA alone, PKA was localized throughout the cytoplasm and yielded a di ffuse staining pattern. Taken together, our results suggest that the direct inhibition of PKA by caveolin-1 is an important and previously unrecognize d mechanism for modulating cAMP-mediated signaling.