title: cGMP-dependent protein kinase I gamma encodes a nuclear localization signal that regulates nuclear compartmentation and function.
abstract: 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 PKGIγ, a proteolytically cleaved PKGI kinase fragment that translocates to the nucleus of SMC. Immuno-localization studies using cells expressing native and NLS-mutant PKGIγ, and treated with a small molecule nuclear transport inhibitor, indicated that PKGIγ encodes a constitutively active NLS that requires importin α and β 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 PKGIγ nuclear function. In addition, although cytosolic PKGIγ-activity was observed to stimulate MAPK/ERK-mediated nuclear CREB signaling in SMC, NLS-mediated PKGIγ nuclear activity alone was determined to increase the expression of differentiation marker proteins in these cells. These results indicate that NLS-mediated nuclear PKGIγ localization plays an important role in how PKGI regulates vascular SMC phenotype.
authors.affiliation: Cardiovascular Research Center, Massachusetts General Hospital, Boston, MA, USA; Harvard Medical School, Cambridge, MA, USA.
authors.forename: Jingsi
authors.initials: J
authors.lastname: Chen
authors.affiliation: Cardiovascular Research Center, Massachusetts General Hospital, Boston, MA, USA; Harvard Medical School, Cambridge, MA, USA; Departments of Anesthesia, Pediatrics, Massachusetts General Hospital, Boston, MA, USA; Department of Medicine, Massachusetts General Hospital, Boston, MA, USA. Electronic address: roberts@cvrc.mgh.harvard.edu.
authors.forename: Jesse D
authors.initials: JD
authors.lastname: Roberts
mesh_terms: D000818:Animals; D001692:Biological Transport / Q000502:physiology; D002454:Cell Differentiation / Q000502:physiology; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D062965:Cyclic GMP-Dependent Protein Kinase Type I / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000502:physiology; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000378:metabolism; D032389:Myocytes, Smooth Muscle / Q000378:metabolism; D019913:Nuclear Localization Signals / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism; D051381:Rats
chemical_list: D017362:Cyclic AMP Response Element-Binding Protein; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins; D062965:Cyclic GMP-Dependent Protein Kinase Type I; D020928:Mitogen-Activated Protein Kinases
keywords: Nuclear translocation; Smooth muscle cell; cGMP signal transduction; cGMP-dependent protein kinase