title: Distinct structural features of G protein-coupled receptor kinase 5 (GRK5) regulate its nuclear localization and DNA-binding ability.
abstract: G protein-coupled receptor kinases (GRKs) act to desensitize G protein-coupled receptors (GPCRs). In addition to this role at the plasma membrane, a nuclear function for GRK5, a member of the GRK4 subfamily of GRKs, has been reported. GRK5 phosphorylates and promotes the nuclear export of the histone deacetylase, HDAC5. Here we demonstrate that the possession of a nuclear localization sequence (NLS) is a common feature of GRK4 subfamily members (GRKs 4, 5 and 6). However, the location of the NLS and the ability of these GRKs to bind DNA in vitro are different. The NLSs of GRK5 and 6 bind DNA in vitro, whilst the NLS of GRK4 does not. Using mutants of GRK5 we identify the regions of GRK5 required for DNA-binding in vitro and nuclear localization in cells. The DNA-binding ability of GRK5 requires both the NLS and an N-terminal calmodulin (CaM)-binding site. A functional nuclear export sequence (NES), required for CaM-dependent nuclear export of the kinase, is also identified. Based on our observations we propose a model to explain how nuclear localization of GRK5 may be regulated. Notably, the nuclear localization of GRK5 and 6 is differentially regulated. These results suggest subfamily specific nuclear functions for the GRK4 subfamily members. Identification of GRK specific small molecule inhibitors of nuclear localization and/or function for the GRK4 subfamily may thus be an achievable goal.
authors: Medical Research Council Laboratory for Molecular Cell Biology and Department of Neuroscience, Physiology and Pharmacology, University College London, London, United Kingdom.
authors: Laura R
authors: LR
authors: Johnson
authors: James D
authors: JD
authors: Robinson
authors: Katrina N
authors: KN
authors: Lester
authors: Julie A
authors: JA
authors: Pitcher
mesh_terms: D021581:Active Transport, Cell Nucleus; D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D004247:DNA / Q000378:metabolism*; D054774:G-Protein-Coupled Receptor Kinase 5 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D019913:Nuclear Localization Signals; D011485:Protein Binding; D061987:Sf9 Cells; D018411:Spodoptera
chemical_list: D019913:Nuclear Localization Signals; D004247:DNA; D054774:G-Protein-Coupled Receptor Kinase 5; D002118:Calcium
references: FEBS Lett. 2004 May 21;566(1-3):275-80
references: 15147908
references: Genomics. 1999 Sep 1;60(2):199-209
references: 10486211
references: J Biol Chem. 2001 May 4;276(18):15192-9
references: 11278469
references: Proc Natl Acad Sci U S A. 2008 Aug 26;105(34):12457-62
references: 18711143
references: Mol Biol Cell. 2007 Aug;18(8):2960-9
references: 17538017
references: J Biol Chem. 1996 Mar 15;271(11):6403-10
references: 8626439
references: J Biol Chem. 2006 Jun 16;281(24):16785-93
references: 16613860
references: EMBO J. 1998 Jun 15;17(12):3363-71
references: 9628873
references: EMBO Rep. 2000 Nov;1(5):411-5
references: 11258480
references: J Biol Chem. 2001 Apr 20;276(16):13015-24
references: 11115515
references: J Biol Chem. 1994 Mar 4;269(9):6832-41
references: 8120045
references: Nat Rev Mol Cell Biol. 2002 Sep;3(9):639-50
references: 12209124
references: Hypertension. 2010 Oct;56(4):696-704
references: 20660817
references: Mol Endocrinol. 1992 Nov;6(11):1867-73
references: 1480176
references: Mol Cell Biol. 2004 Dec;24(23):10169-79
references: 15542828
references: Biochem J. 2009 Dec 14;425(1):169-78
references: 19796012
references: J Biol Chem. 1996 Oct 4;271(40):24907-13
references: 8798768
references: Cell Signal. 2005 Aug;17(8):917-28
references: 15894165
references: Proc Natl Acad Sci U S A. 2008 Nov 18;105(46):17818-23
references: 19008357
references: J Biol Chem. 1997 Jul 18;272(29):18273-80
references: 9218466
references: Science. 2003 May 23;300(5623):1256-62
references: 12764189
references: J Mol Med (Berl). 2009 May;87(5):455-63
references: 19229505
references: Mol Cell Biol. 1999 Jan;19(1):690-703
references: 9858592
references: Cell Signal. 2003 Nov;15(11):973-81
references: 14499340
references: Biochim Biophys Acta. 2007 Apr;1768(4):913-22
references: 17084806
references: J Biol Chem. 1999 Oct 8;274(41):29381-9
references: 10506199
references: J Biol Chem. 1994 Apr 8;269(14):10209-12
references: 8144599
references: Nucleic Acids Res. 2003 Jan 1;31(1):393-6
references: 12520031