Association of alphaB-crystallin, a small heat shock protein, with actin: role in modulating actin filament dynamics in vivo.
Disruption of cytoskeletal assembly is one of the early effects of any stress that can ultimately lead to cell death. Stabilization of cytoskeletal assembly, therefore, is a critical event that regulates cell survival under stress. alphaB-crystallin, a small heat shock protein, has been shown to associate with cytoskeletal proteins under normal and stress conditions. Earlier reports suggest that alphaB-crystallin could prevent stress-induced aggregation of actin in vitro. However, the molecular mechanisms by which alphaB-crystallin stabilizes actin filaments in vivo are not known. Using the H9C2 rat cardiomyoblast cell line as a model system, we show that upon heat stress, alphaB-crystallin preferentially partitions from the soluble cytosolic fraction to the insoluble cytoskeletal protein-rich fraction. Confocal microscopic analysis shows that alphaB-crystallin associates with actin filaments during heat stress and the extent of association increases with time. Further, immunoprecipitation experiments show that alphaB-crystallin interacts directly with actin. Treatment of heat-stressed H9C2 cells with the actin depolymerzing agent, cytochalasin B, failed to disorganize actin. We show that this association of alphaB-crystallin with actin is dependent on its phosphorylation status, as treatment of cells with MAPK inhibitors SB202190 or PD98059 results in abrogation of this association. Our results indicate that alphaB-crystallin regulates actin filament dynamics in vivo and protects cells from stress-induced death. Further, our studies suggest that the association of alphaB-crystallin with actin helps maintenance of pinocytosis, a physiological function essential for survival of cells.
Singh
Bhairab N
BN
Centre for Cellular and Molecular Biology, Hyderabad, India.
Rao
K Sridhar
KS
Ramakrishna
Tangirala
T
Rangaraj
Nandini
N
Rao
Ch Mohan
ChM
D008841:Actin Cytoskeleton / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D000199:Actins / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D003571:Cytochalasin B / Q000494:pharmacology; D003911:Dextrans / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016650:Fluorescein-5-isothiocyanate; D050888:Heat-Shock Proteins, Small / Q000378:metabolism*; D018869:Heat-Shock Response / Q000502:physiology; D006358:Hot Temperature; D006979:Hyperthermia, Induced; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors; D010766:Phosphorylation / Q000187:drug effects; D010873:Pinocytosis / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D051381:Rats; D012995:Solubility / Q000187:drug effects; D038203:alpha-Crystallin B Chain / Q000378:metabolism*
D000199:Actins; D003911:Dextrans; D004791:Enzyme Inhibitors; D050888:Heat-Shock Proteins, Small; D038203:alpha-Crystallin B Chain; D003571:Cytochalasin B; D020928:Mitogen-Activated Protein Kinases; D016650:Fluorescein-5-isothiocyanate