Yeast Rsp5 ubiquitin ligase affects the actin cytoskeleton in vivo and in vitro

Eur J Cell Biol. 2011 Dec;90(12):1016-28. doi: 10.1016/j.ejcb.2011.08.002. Epub 2011 Oct 14.


Yeast Rsp5 ubiquitin ligase is involved in several cellular processes, including endocytosis. Actin patches are sites of endocytosis, a process involving actin assembly and disassembly. Here we show Rsp5 localization in cortical patches and demonstrate its involvement in actin cytoskeleton organization and dynamics. We found that the Rsp5-F1-GFP2 N-terminal fragment and full length GFP-Rsp5 were recruited to peripheral patches that temporarily co-localized with Abp1-mCherry, a marker of actin patches. Actin cytoskeleton organization was defective in a strain lacking RSP5 or overexpressing RSP5, and this phenotype was accompanied by morphological abnormalities. Overexpression of RSP5 caused hypersensitivity of cells to Latrunculin A, an actin-depolymerizing drug and was toxic to cells lacking Las17, an activator of actin nucleation. Moreover, Rsp5 was required for efficient actin polymerization in a whole cell extract based in vitro system. Rsp5 interacted with Las17 and Las17-binding proteins, Lsb1 and Lsb2, in a GST-Rsp5-WW2/3 pull down assay. Rsp5 ubiquitinated Lsb1-HA and Lsb2-HA without directing them for degradation. Overexpression of RSP5 increased the cellular level of HA-Las17 in wild type and in lsb1Δ lsb2Δ strains in which the basal level of Las17 was already elevated. This increase was prevented in a strain devoid of Las17-binding protein Sla1 which is also a target of Rsp5 ubiquitination. Thus, Rsp5 together with Lsb1, Lsb2 and Sla1 regulate the level of Las17, an important activator of actin polymerization.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Actin Cytoskeleton / genetics
  • Actin Cytoskeleton / metabolism*
  • Cytoskeleton / enzymology
  • Cytoskeleton / metabolism*
  • Endosomal Sorting Complexes Required for Transport / genetics
  • Endosomal Sorting Complexes Required for Transport / metabolism*
  • Gene Expression Regulation, Fungal
  • Humans
  • Saccharomyces cerevisiae / drug effects
  • Saccharomyces cerevisiae / enzymology*
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism*
  • Ubiquitin-Protein Ligase Complexes / genetics
  • Ubiquitin-Protein Ligase Complexes / metabolism*
  • Wiskott-Aldrich Syndrome Protein / genetics
  • Wiskott-Aldrich Syndrome Protein / metabolism


  • Endosomal Sorting Complexes Required for Transport
  • LAS17 protein, S cerevisiae
  • Saccharomyces cerevisiae Proteins
  • Wiskott-Aldrich Syndrome Protein
  • Ubiquitin-Protein Ligase Complexes
  • RSP5 protein, S cerevisiae