An actin-regulated importin α/β-dependent extended bipartite NLS directs nuclear import of MRTF-A.
Myocardin-related transcription factors (MRTFs) are actin-regulated transcriptional coactivators, which bind G-actin through their N-terminal RPEL domains. In response to signal-induced actin polymerisation and concomitant G-actin depletion, MRTFs accumulate in the nucleus and activate target gene transcription through their partner protein SRF. Nuclear accumulation of MRTFs in response to signal is inhibited by increased G-actin level. Here, we study the mechanism by which MRTF-A enters the nucleus. We show that MRTF-A contains an unusually long bipartite nuclear localisation signal (NLS), comprising two basic elements separated by 30 residues, embedded within the RPEL domain. Using siRNA-mediated protein depletion in vivo, and nuclear import assays in vitro, we show that the MRTF-A extended bipartite NLS uses the importin (Imp)α/β-dependent import pathway, and that import is inhibited by G-actin. Interaction of the NLS with the Impα-Impβ heterodimer requires both NLS basic elements, and is dependent on the Impα major and minor binding pockets. Binding of the Impα-Impβ heterodimer to the intact MRTF-A RPEL domain occurs competitively with G-actin. Thus, MRTF-A contains an actin-sensitive nuclear import signal.
Pawłowski
Rafał
R
Transcription Laboratory, Cancer Research UK, London Research Institute, London, UK.
Rajakylä
Eeva Kaisa
EK
Vartiainen
Maria K
MK
Treisman
Richard
R
D000199:Actins / Q000378:metabolism*; D021581:Active Transport, Cell Nucleus / Q000502:physiology*; D000595:Amino Acid Sequence; D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D041681:NIH 3T3 Cells; D019913:Nuclear Localization Signals / Q000235:genetics* / Q000378:metabolism*; D034622:RNA Interference; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D028901:alpha Karyopherins / Q000235:genetics / Q000378:metabolism*; D028961:beta Karyopherins / Q000235:genetics / Q000378:metabolism*
D000199:Actins; C511173:Mrtfa protein, mouse; D019913:Nuclear Localization Signals; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; D028901:alpha Karyopherins; D028961:beta Karyopherins
J Cell Sci. 2010 Jan 1;123(Pt 1):23-8
20016063
Mol Cell Biol. 2003 Sep;23(18):6597-608
12944485
Mol Cell Biol. 2004 Dec;24(23):10246-55
15542834
Cell Physiol Biochem. 2002;12(5-6):335-44
12438769
Traffic. 2005 Mar;6(3):187-98
15702987
Trends Cell Biol. 2004 Sep;14(9):505-14
15350979
Cell. 1998 Jul 24;94(2):193-204
9695948
Annu Rev Cell Dev Biol. 1999;15:607-60
10611974
EMBO J. 2004 Oct 13;23(20):3973-83
15385960
Cell Mol Life Sci. 2003 Aug;60(8):1659-88
14504656
Nat Cell Biol. 2000 Jun;2(6):376-8
10854330
Mol Cell Biol. 1999 Nov;19(11):7782-91
10523667
Structure. 2000 Mar 15;8(3):329-38
10745017
J Mol Biol. 2000 Apr 14;297(5):1183-94
10764582
J Biol Chem. 2005 Apr 22;280(16):15942-51
15677444
Nat Struct Mol Biol. 2007 May;14(5):452-4
17435768
Nat Methods. 2006 Dec;3(12):995-1000
17072308
J Biol Chem. 2007 Feb 23;282(8):5101-5
17170104
Nat Cell Biol. 2007 Jan;9(1):72-9
17159997
Nat Cell Biol. 2009 Mar;11(3):257-68
19198601
Cell. 1991 Feb 8;64(3):615-23
1991323
Mol Genet Genomics. 2002 Nov;268(3):287-97
12436251
Genes Dev. 1992 Oct;6(10):1899-913
1340770
Genes Dev. 1998 Sep 1;12(17):2673-83
9732266
J Mol Evol. 2009 Apr;68(4):351-65
19308634
Methods Enzymol. 2002;351:587-607
12073370
J Biol Chem. 1997 Feb 14;272(7):4310-5
9020149
J Biol Chem. 2009 Jan 2;284(1):478-485
19001369
Annu Rev Cell Dev Biol. 1999;15:291-339
10611964
Oncogene. 2003 Feb 20;22(7):1057-69
12592392
Cell. 2003 May 2;113(3):329-42
12732141
Cell. 2004 May 28;117(5):611-23
15163409
EMBO J. 2008 Dec 3;27(23):3198-208
19008859
Mol Cell Biol. 2008 Jan;28(2):732-42
18025109
Curr Biol. 1995 Apr 1;5(4):383-92
7627554
Trends Biochem Sci. 1991 Dec;16(12):478-81
1664152
Biochim Biophys Acta. 2008 Mar;1783(3):394-404
18187047
J Biol Chem. 2003 Jul 25;278(30):27981-7
12695505
Chem Biol. 2008 Sep 22;15(9):940-9
18804031
Nat Rev Mol Cell Biol. 2007 Mar;8(3):195-208
17287812
Traffic. 2005 Mar;6(3):173-86
15702986
Cell. 1999 Jul 23;98(2):159-69
10428028
Science. 2007 Jun 22;316(5832):1749-52
17588931
Curr HIV Res. 2010 Jan;8(1):2-13
20210777
Traffic. 2010 Mar;11(3):311-23
20028483