Huntingtin associates with the actin cytoskeleton and α-actinin isoforms to influence stimulus dependent morphology changes

PLoS One. 2019 Feb 15;14(2):e0212337. doi: 10.1371/journal.pone.0212337. eCollection 2019.

Abstract

One response of cells to growth factor stimulus involves changes in morphology driven by the actin cytoskeleton and actin associated proteins which regulate functions such as cell adhesion, motility and in neurons, synaptic plasticity. Previous studies suggest that Huntingtin may be involved in regulating morphology however, there has been limited evidence linking endogenous Huntingtin localization or function with cytoplasmic actin in cells. We found that depletion of Huntingtin in human fibroblasts reduced adhesion and altered morphology and these phenotypes were made worse with growth factor stimulation, whereas the presence of the Huntington's Disease mutation inhibited growth factor induced changes in morphology and increased numbers of vinculin-positive focal adhesions. Huntingtin immunoreactivity localized to actin stress fibers, vinculin-positive adhesion contacts and membrane ruffles in fibroblasts. Interactome data from others has shown that Huntingtin can associate with α-actinin isoforms which bind actin filaments. Mapping studies using a cDNA encoding α-actinin-2 showed that it interacts within Huntingtin aa 399-969. Double-label immunofluorescence showed Huntingtin and α-actinin-1 co-localized to stress fibers, membrane ruffles and lamellar protrusions in fibroblasts. Proximity ligation assays confirmed a close molecular interaction between Huntingtin and α-actinin-1 in human fibroblasts and neurons. Huntingtin silencing with siRNA in fibroblasts blocked the recruitment of α-actinin-1 to membrane foci. These studies support the idea that Huntingtin is involved in regulating adhesion and actin dependent functions including those involving α-actinin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Actin Cytoskeleton / chemistry
  • Actin Cytoskeleton / metabolism*
  • Actinin / metabolism*
  • Cell Adhesion / drug effects
  • Cell Membrane / metabolism
  • Cells, Cultured
  • Fibroblasts / cytology
  • Fibroblasts / pathology
  • Humans
  • Huntingtin Protein / antagonists & inhibitors
  • Huntingtin Protein / genetics
  • Huntingtin Protein / metabolism*
  • Huntington Disease / metabolism
  • Huntington Disease / pathology
  • Microscopy, Confocal
  • Neurons / metabolism
  • Neurons / pathology
  • Platelet-Derived Growth Factor / pharmacology
  • Protein Isoforms / metabolism
  • RNA Interference
  • RNA, Small Interfering / metabolism

Substances

  • ACTN1 protein, human
  • HTT protein, human
  • Huntingtin Protein
  • Platelet-Derived Growth Factor
  • Protein Isoforms
  • RNA, Small Interfering
  • Actinin

Grant support

This work was supported by contracts from CHDI, Inc. to KKG and MD and the Dake Family Fund to MD.