title: cGMP-dependent protein kinase anchoring by IRAG regulates its nuclear translocation and transcriptional activity.
abstract: Type I cGMP-dependent protein kinases (PKGs) translocate to the nucleus to regulate gene expression in some, but not all cell types; we hypothesized that nuclear translocation of PKG may be regulated by extra-nuclear anchoring proteins. The inositol 1,4,5-triphosphate (IP(3)) receptor-associated cGMP kinase substrate (IRAG) binds to the N-terminus of PKG Ibeta, but not PKG Ialpha, and in smooth muscle cells, IRAG and PKG Ibeta are in a complex with the IP(3) receptor at endoplasmatic reticulum membranes, where the complex regulates calcium release [Schlossmann et al., Nature, 404 (2000) 197]. We found that co-expression of IRAG and PKG Ibeta in baby hamster kidney cells prevented cGMP-induced PKG Ibeta translocation to the nucleus, and decreased cGMP/PKG Ibeta transactivation of a cAMP-response element-dependent reporter gene. These effects required the PKG Ibeta/IRAG association, as demonstrated by a binding-incompetent IRAG mutant, and were specific for PKG Ibeta, as nuclear translocation and reporter gene activation by PKG Ialpha was not affected by IRAG. A phosphorylation-deficient IRAG mutant that is no longer functionally regulated by PKG phosphorylation suppressed cGMP/PKG Ibeta transcriptional activity, indicating that IRAG's effect was not explained by changes in intracellular calcium, and was not related to competition of IRAG with other PKG substrates. These results demonstrate that PKG anchoring to a specific binding protein is sufficient to dictate subcellular localization of the kinase and affect cGMP signaling in the nucleus, and may explain why nuclear translocation of PKG I does not occur in all cell types.
authors.affiliation: Department of Medicine and Cancer Center, University of California at San Diego, La Jolla, CA 92093-0652, United States.
authors.forename: Darren E
authors.initials: DE
authors.lastname: Casteel
authors.forename: Tong
authors.initials: T
authors.lastname: Zhang
authors.forename: Shunhui
authors.initials: S
authors.lastname: Zhuang
authors.forename: Renate B
authors.initials: RB
authors.lastname: Pilz
mesh_terms: D000818:Animals; D002460:Cell Line; D002467:Cell Nucleus / Q000201:enzymology*; D017869:Cyclic GMP-Dependent Protein Kinases / Q000235:genetics* / Q000378:metabolism*; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D007527:Isoenzymes / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D050505:Mutant Proteins / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D021381:Protein Transport; D013347:Subcellular Fractions / Q000201:enzymology; D014158:Transcription, Genetic; D015533:Transcriptional Activation
chemical_list: D007527:Isoenzymes; D008565:Membrane Proteins; D050505:Mutant Proteins; D017869:Cyclic GMP-Dependent Protein Kinases