Distribution, phosphorylation, and activities of Hsp25 in heat-stressed H9c2 myoblasts: a functional link to cytoprotection

Abstract

The behavior of the endogenous heat shock protein 25 (Hsp25) in heat-stressed rat H9c2 myoblasts was studied. After mild or severe heating, this protein became less extractable with Triton X-100 and displayed characteristic immunofluorescence patterns, namely (1) granules in the nucleus, and (2) association with F-actin bundles in the cytoplasm. The intranuclear granulation of Hsp25 and its association with F-actin were sensitive to drugs affecting Hsp25 phosphorylation (cantharidin, sodium orthovanadate, SB203580, SB202190). Isoform analysis of Hsp25 translocated to the nucleus-free cytoskeletal fraction revealed only mono- and biphosphorylated Hsp25 and no unphosphorylated Hsp25. Transfected luciferase with initial localization in the nucleosol became colocalized with the Hsp25-containing granules after a heat shock treatment that denatured the enzyme in the cells. The association of Hsp25 with actin filaments after a mild heat stress conferred protection from subsequent F-actin-damaging treatments with cytochalasins (D and B) or severe heat stress. We hypothesize that (1) the binding of heat-denatured nucleosolic proteins to the Hsp25 contained in specific granular structures may serve for the subsequent chaperoning or degradation of the bound proteins, and (2) the actin cytoskeleton is stabilized by the direct targeting of phosphorylated Hsp25 to microfilament bundles.

In vitro evaluation of aspirin-induced HspB1 against heat stress damage in chicken myocardial cells

“…It is localized within the perinuclear region at normal conditions (37°C) and is relocated to the nucleus after heat stress (Arrigo et al 1988). Stress from heat or ATP-depletion causes HspB1 to form large (∼106 kDa) detergent insoluble structures inside the nucleus (Arrigo et al 1988;Arrigo 1994), which are visible as granules (Adhikari et al 2004;Bryantsev et al 2002;Loktionova et al 1996) and also contain heat-denatured proteins. Therefore, the protective roles of HspB1 are also regulated by phosphorylation and translocation.…”

Recruitment of phosphorylated small heat shock protein Hsp27 to nuclear speckles without stress

“…Despite consequent interest in cellular roles played by Hsp27, neither the mechanism nor function of Hsp27 are fully understood. Experimental evidence supports a variety of roles for Hsp27, including a general chaperone activity [1,5,6], interaction with actin cytoskeleton elements [7][8][9][10], interaction with pro-and anti-apoptotic signaling factors [11][12][13], and modulation of the oxidative balance of cells [14]. These activities can occur throughout the cell or are restricted to the cytoplasmic cell compartment.…”

HSP25 Affects the Proliferation and Differentiation of Rat Dental Follicle Cells

“…In response to stress, HSP25 will translocate in two ways: ①The cytosolic pool of this protein migrates to the nucleus to combine with the denatured proteins, and may function in protecting the cell from being killed. ②HSP25 linked to F-actin bundles, enhances the resistance to cytoskeleton-damaging insults (Bryantsev et al, 2002). We may expect that HSP25 will protect DFCs against outside stress in similar ways.…”