title: An actin-regulated importin α/β-dependent extended bipartite NLS directs nuclear import of MRTF-A.
abstract: 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.
authors.affiliation: Transcription Laboratory, Cancer Research UK, London Research Institute, London, UK.
authors.forename: Rafał
authors.initials: R
authors.lastname: Pawłowski
authors.forename: Eeva Kaisa
authors.initials: EK
authors.lastname: Rajakylä
authors.forename: Maria K
authors.initials: MK
authors.lastname: Vartiainen
authors.forename: Richard
authors.initials: R
authors.lastname: Treisman
mesh_terms: 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*
chemical_list: D000199:Actins; C511173:Mrtfa protein, mouse; D019913:Nuclear Localization Signals; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; D028901:alpha Karyopherins; D028961:beta Karyopherins
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