Small heat shock protein Hsp27 prevents heat‐induced aggregation of F‐actin by forming soluble complexes with denatured actin

Abstract

Previously, we have shown that the small heat shock protein with apparent molecular mass 27 kDa (Hsp27) does not affect the thermal unfolding of F‐actin, but effectively prevents aggregation of thermally denatured F‐actin [Pivovarova AV, Mikhailova VV, Chernik IS, Chebotareva NA, Levitsky DI & Gusev NB (2005) Biochem Biophys Res Commun331, 1548–1553], and supposed that Hsp27 prevents heat‐induced aggregation of F‐actin by forming soluble complexes with denatured actin. In the present work, we applied dynamic light scattering, analytical ultracentrifugation and size exclusion chromatography to examine the properties of complexes formed by denatured actin with a recombinant human Hsp27 mutant (Hsp27–3D) mimicking the naturally occurring phosphorylation of this protein at Ser15, Ser78, and Ser82. Our results show that formation of these complexes occurs upon heating and accompanies the F‐actin thermal denaturation. All the methods show that the size of actin–Hsp27‐3D complexes decreases with increasing Hsp27‐3D concentration in the incubation mixture and that saturation occurs at approximately equimolar concentrations of Hsp27‐3D and actin. Under these conditions, the complexes exhibit a hydrodynamic radius of ∼ 16 nm, a sedimentation coefficient of 17–20 S, and a molecular mass of about 2 MDa. It is supposed that Hsp27‐3D binds to denatured actin monomers or short oligomers dissociated from actin filaments upon heating and protects them from aggregation by forming relatively small and highly soluble complexes. This mechanism might explain how small heat shock proteins prevent aggregation of denatured actin and by this means protect the cytoskeleton and the whole cell from damage caused by accumulation of large insoluble aggregates under heat shock conditions.

Gamma Interferon-Induced Guanylate Binding Protein 1 Is a Novel Actin Cytoskeleton Remodeling Factor

“…Hydrodynamic radii derived from DLS measurements have previously been used to determine the size and shape of proteins (63,64). Actin polymerization was initiated again in the presence or absence of GBP-1, but in contrast to AFM, the DLS method determined fragment length directly in solution.…”

Roles of the N‐ and C‐terminal sequences in Hsp27 self‐association and chaperone activity

“…For example, Hsp27 exhibits chaperone activity, 2 inhibits cytochrome c-induced apoptosis by inhibiting activation of procaspase-9, 3 modulates oxidative stress and regulates the cytoskeleton. [4][5][6][7][8] In addition, overexpression of Hsp27 by tumor cells increases their tumorigenicity and protects against cell death triggered by a number of stimuli, e.g., hyperthermia, oxidative stress, staurosporine, ligation of the Fas/ApoÀ1/CD95 death receptor, and cytotoxic drugs. 9 Mutant forms of Hsp27 have been linked to human neuromuscular disorders such as axonal CharcotÀMarieÀTooth disease 10,11 and distal hereditary motor neuropathy, 12 and Hsp27 accumulates in individuals with various neurodegenerative disorders, including Alzheimer's, Parkinson's, Alexander's, and CreutzfeldtÀJakob diseases and multiple sclerosis.…”

Regulation of Endothelial Permeability by Glutathione S-Transferase Pi Against Actin Polymerization

“…As a small molecular chaperone, HSP27 interacts with F-actin as a monomer. When HSP27 is phosphorylated, it dissociates from the positive end of F-actin, which leads to intracellular F-actin depolymerization and remodeling and the formation of stress fibers, resulting in increased cell permeability [21]. We further analyzed the effects of GSTpi on the p38MAPK/MK2/HSP27 signaling pathway.…”