Identification of alpha-tubulin as an hsp105alpha-binding protein by the yeast two-hybrid system

Exp Cell Res. 2003 Jun 10;286(2):233-40. doi: 10.1016/s0014-4827(03)00054-5.

Abstract

Hsp105alpha is a mammalian stress protein that belongs to the HSP105/110 family. Hsp105alpha prevents stress-induced apoptosis in neuronal cells and binds to Hsp70/Hsc70 and suppresses the Hsp70 chaperone activity in vitro. In this study, to further elucidate the function of Hsp105alpha, we searched for Hsp105alpha-binding proteins by screening a mouse FM3A cell cDNA library with full-length Hsp105alpha using the yeast two-hybrid system and obtained alpha-tubulin as an Hsp105alpha-binding protein. Hsp105alpha bound directly to alpha-tubulin both in vitro and in vivo. Indirect immunofluorescence analysis with anti-Hsp105 and anti-alpha-tubulin antibodies indicated that Hsp105alpha was colocalized with microtubules. Furthermore, the disorganization of microtubules induced by heat shock was prevented in Hsp105alpha-overexpressing COS-7 cells. These findings suggested that Hsp105alpha associates with alpha-tubulin and microtubules in cells and plays a role in protection of microtubules under conditions of stress.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Apoptosis / genetics
  • COS Cells
  • DNA, Complementary / analysis
  • DNA, Complementary / genetics
  • Eukaryotic Cells / metabolism*
  • HSP110 Heat-Shock Proteins
  • HSP70 Heat-Shock Proteins / genetics
  • HSP70 Heat-Shock Proteins / metabolism*
  • HeLa Cells
  • Heat-Shock Response / genetics
  • Humans
  • Mice
  • Microtubules / metabolism*
  • Protein Binding / genetics
  • Protein Structure, Tertiary / genetics
  • Recovery of Function / genetics
  • Stress, Physiological / genetics
  • Stress, Physiological / metabolism*
  • Tubulin / genetics
  • Tubulin / metabolism*
  • Two-Hybrid System Techniques

Substances

  • DNA, Complementary
  • HSP110 Heat-Shock Proteins
  • HSP70 Heat-Shock Proteins
  • HSPH1 protein, human
  • Hsp105 protein, mouse
  • Tubulin