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In response to stress, the expression level of HSP27 increases several fold to confer cellular resistance to the adverse environmental change. HSP27 is localized to the cytoplasm of unstressed cells but can redistribute to the nucleus in response to stress, where it may function to stabilize DNA and/or the nuclear membrane. HSP27 is phosphorylated at serines 15, 78 and 82 by MAPKAPK-2 as a result of the activation of the p38 MAPK pathway. Phosphorylation of HSP27 causes a change in its tertiary structure, which shifts from large homotypic multimers to dimers and monomers. Thus, phosphorylated HSPs have significantly decreased ability to act as molecular chaperones. Phosphorylation of HSP27 on Ser82 by MAPKAPK-2 or AKT leads to HSP dissociation from the AKT/MAPKAPK2/p38 complex, which may promote independent survival signals. The phosphorylation of HSP27 also enhances its association with IKKβ to result in decreased IKK activity.
Figure 1: Measurement of phosphorylated HSP27 (S82) and total HSP27.
Human HeLa cells were cultured in 96-well plates and serum-starved (0.5% FBS) for 16 hours. Cells were then stimulated with 25 µg/ml of Anisomycin for 30 minutes and fixed. Total HSP27 and HSP27 phosphorylated on Ser82 were each assayed in duplicate using the phospho-HSP27 (S82) and total-HSP27 antibodies included in the FACE HSP27 Kit. Data were plotted after correction for cell number (performed through use of Crystal Violet).
Antibody Specificities
The phospho-HSP27 (Ser82) antibody was raised against a synthetic phospho-peptide corresponding to residues surrounding phosphorylated Ser 82 of human HSP27. The phospho-HSP27 (Ser82) antibody recognizes HSP27 only when phosphorylated at Ser82. The total-HSP27 antibody recognizes HSP27 protein regardless of its phosphorylation state.
