cGMP-dependent protein kinase I gamma encodes a nuclear localization signal that regulates nuclear compartmentation and function

cGMP-dependent protein kinase I (PKGI) plays an important role in regulating how cGMP specifies vascular smooth muscle cell (SMC) phenotype. Although studies indicate that PKGI nuclear localization controls how cGMP regulates gene expression in SMC, information about the mechanisms that regulate PKGI nuclear compartmentation and its role in directly regulating cell phenotype is limited. Here we characterize a nuclear localization signal sequence (NLS) in PKGI gamma, a proteolytically cleaved PKGI kinase fragment that translocates to the nucleus of SMC Immuno-localization studies using cells expressing native and NLS-mutant PKGl gamma, and treated with a small molecule nuclear transport inhibitor, indicated that PKGl gamma encodes a constitutively active NLS that requires importin alpha and beta for regulation of its compartmentation. Moreover, studies utilizing a genetically encoded nuclear phospho-CREB biosensor probe and fluorescence lifetime imaging microscopy demonstrated that this NLS controls PKGl gamma nuclear function. In addition, although cytosolic PKGl gamma-activity was observed to stimulate MAPK/ERK-mediated nuclear CREB signaling in SMC, NLS-mediated PKGI gamma nuclear activity alone was determined to increase the expression of differentiation marker proteins in these cells. These results indicate that NLS-mediated nuclear PKGl gamma localization plays an important role in how PKGI regulates vascular SMC phenotype. (C) 2014 Elsevier Inc. All rights reserved.