An actin-regulated importin α/β-dependent extended bipartite NLS directs nuclear import of MRTF-A

EMBO J. 2010 Oct 20;29(20):3448-58. doi: 10.1038/emboj.2010.216. Epub 2010 Sep 3.

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Actins / metabolism*
  • Active Transport, Cell Nucleus / physiology*
  • Amino Acid Sequence
  • Animals
  • Cell Nucleus / metabolism
  • Mice
  • Molecular Sequence Data
  • NIH 3T3 Cells
  • Nuclear Localization Signals / genetics*
  • Nuclear Localization Signals / metabolism*
  • RNA Interference
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Trans-Activators / genetics
  • Trans-Activators / metabolism*
  • alpha Karyopherins / genetics
  • alpha Karyopherins / metabolism*
  • beta Karyopherins / genetics
  • beta Karyopherins / metabolism*

Substances

  • Actins
  • Mrtfa protein, mouse
  • Nuclear Localization Signals
  • Recombinant Fusion Proteins
  • Trans-Activators
  • alpha Karyopherins
  • beta Karyopherins