CYCLIC NUCLEOTIDE-DEPENDENT VASORELAXATION IS ASSOCIATED WITH THE PHOSPHORYLATION OF A SMALL HEAT SHOCK-RELATED PROTEIN

Activation of cyclic nucleotide-dependent signaling pathways leads to the relaxation of various smooth muscles. One of the major phosphoryla tion events associated with cyclic nucleotide-dependent vasorelaxation in bovine trachealis and carotid artery smooth muscle is the phosphor ylation of two 20-kDa phosphoproteins with pI values of 5.7 and 5.9 (p reviously designated pp8 and pp3, respectively). The present studies s ought to determine the identities of pp3 and pp8 in vascular smooth mu scle. The phosphopeptide maps for the pp8 and pp3 proteins were simila r. Preparative two-dimensional gel electrophoresis and amino acid sequ encing of a peptide fragment of the pp3 protein revealed a sequence id entical to a 20-kDa heat shock-related protein (HSP20) previously puri fied from skeletal muscle. Western blot and immunoprecipitation analys is with anti-HSP20 antibodies demonstrated that the pp3 and pp8 protei ns are phosphorylated forms of HSP20. In addition, HSP20 could be phos phorylated in vitro by both cAMP-dependent protein kinase and cGMP-dep endent protein kinase, These data suggest that the phosphorylation of the heat shock-related protein HSP20 is associated with cyclic nucleot ide-dependent relaxation of vascular smooth muscle.