Non-muscle myosin II regulates neuronal actin dynamics by interacting with guanine nucleotide exchange factors

PLoS One. 2014 Apr 21;9(4):e95212. doi: 10.1371/journal.pone.0095212. eCollection 2014.

Abstract

Background: Non-muscle myosin II (NM II) regulates a wide range of cellular functions, including neuronal differentiation, which requires precise spatio-temporal activation of Rho GTPases. The molecular mechanism underlying the NM II-mediated activation of Rho GTPases is poorly understood. The present study explored the possibility that NM II regulates neuronal differentiation, particularly morphological changes in growth cones and the distal axon, through guanine nucleotide exchange factors (GEFs) of the Dbl family.

Principal findings: NM II colocalized with GEFs, such as βPIX, kalirin and intersectin, in growth cones. Inactivation of NM II by blebbistatin (BBS) led to the increased formation of short and thick filopodial actin structures at the periphery of growth cones. In line with these observations, FRET analysis revealed enhanced Cdc42 activity in BBS-treated growth cones. BBS treatment also induced aberrant targeting of various GEFs to the distal axon where GEFs were seldom observed under physiological conditions. As a result, numerous protrusions and branches were generated on the shaft of the distal axon. The disruption of the NM II-GEF interactions by overexpression of the DH domains of βPIX or Tiam1, or by βPIX depletion with specific siRNAs inhibited growth cone formation and induced slender axons concomitant with multiple branches in cultured hippocampal neurons. Finally, stimulation with nerve growth factor induced transient dissociation of the NM II-GEF complex, which was closely correlated with the kinetics of Cdc42 and Rac1 activation.

Conclusion: Our results suggest that NM II maintains proper morphology of neuronal growth cones and the distal axon by regulating actin dynamics through the GEF-Rho GTPase signaling pathway.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Actins / metabolism*
  • Animals
  • Axons / drug effects
  • Axons / metabolism
  • Cells, Cultured
  • Female
  • Growth Cones / drug effects
  • Growth Cones / metabolism
  • Heterocyclic Compounds, 4 or More Rings / pharmacology
  • Hippocampus / cytology
  • Models, Biological
  • Nerve Growth Factors / metabolism
  • Neurites / drug effects
  • Neurites / metabolism
  • Neurons / drug effects
  • Neurons / metabolism*
  • Nonmuscle Myosin Type IIB / metabolism*
  • Protein Binding / drug effects
  • Protein Structure, Tertiary
  • Pseudopodia / drug effects
  • Pseudopodia / metabolism
  • Rats, Sprague-Dawley
  • Rho Guanine Nucleotide Exchange Factors / chemistry
  • Rho Guanine Nucleotide Exchange Factors / metabolism*
  • Signal Transduction / drug effects
  • cdc42 GTP-Binding Protein / metabolism
  • p21-Activated Kinases / metabolism
  • rac1 GTP-Binding Protein / metabolism

Substances

  • Actins
  • Heterocyclic Compounds, 4 or More Rings
  • Nerve Growth Factors
  • Rho Guanine Nucleotide Exchange Factors
  • blebbistatin
  • p21-Activated Kinases
  • Nonmuscle Myosin Type IIB
  • cdc42 GTP-Binding Protein
  • rac1 GTP-Binding Protein

Grant support

This work was supported by a grant from the National Research Foundation of Korea (NRF-2010-0021813 and NRF-2007-0054943). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.