Abstract 1557: Compartmentation of Cyclic Nucleotide Signaling via Differential Expression of Adenylyl Cyclase Isoforms
Compartmentation of cyclic nucleotide signaling is critical in the regulation of signaling via adenylyl cyclase activation. This compartmentation has previously been believed to be due to the structural association between phosphodiesterases (PDEs) and PKA (via their association with AKAPs). To determine whether different isoforms of adenylyl cyclase may be differentially compartmentalized and whether this relates to differences in the functional effects of adenylyl cyclase activation we studied the effect of the expression of representative adenylyl cyclase isoforms (AC1, AC2, AC5 and AC6) on cellular cAMP activity, PKA activation, arborization responses (an index of cAMP-mediated cytoskeletal re-organization) as well as ERK activation and cell growth in vascular smooth muscle cells (VSMCs) with adenoviral-AC infections. When isoforms were expressed at levels to comparably enhance forskolin-stimulated AC activity, only gene transfer of AC6 significantly enhanced forskolin-stimulated cAMP accumulation, PKA activation and VASP phosphorylation. In addition, only AC6 gene transfer enhanced the arborization responses in VSMCs (142±15% of GFP-infected VSMCs, n=6). In contrast, following PDE inhibition (using IBMX), the extent of cAMP accumulation, PKA activation and VASP phosphorylation did not differ between isoforms. In regards to regulation of ERK activation and regulation of cell growth, expression of AC1, alone among the isoforms tested, enhanced forskolin-stimulated ERK phosphorylation, paralleling a demonstrable physical association between AC1 and ERK1/2 could be demonstrated by co-immunoprecipitation techniques. Further, AC1 expression inhibits ERK1/2 distribution to nuclear fractions. In addition, AC1 gene transfer increased cell doubling times (176±22% of GFP, n=3) and enhanced forskolin-mediated inhibition of cell growth. These data support a previously unappreciated differences in the association of specific isoforms of AC with PDE (less for AC6 vs. other isoforms tested) as well as differences in their ability to association with ERK (more for AC1) and hence regulation of cellular growth. These differences may be critical in the regulation of cyclic nucleotide-mediated vascular functions.