Cell stress promotes the association of phosphorylated HspB1 with F-actin

Autor: Karen M. Mearow, Joseph P. Clarke
Rok vydání: 2013
Předmět:
HSP27 Heat-Shock Proteins
lcsh:Medicine
PC12 Cells
Biochemistry
chemistry.chemical_compound
0302 clinical medicine
Molecular cell biology
RNA interference
Tissue Distribution
Phosphorylation
Cytoskeleton
lcsh:Science
Cellular localization
Cellular Stress Responses
0303 health sciences
Multidisciplinary
Immunochemistry
Statistics
Cellular Structures
Cell biology
Cytochemistry
Immunocytochemistry
Protein Binding
Research Article
animal structures
Phalloidin
Immunoprecipitation
macromolecular substances
Biology
Biostatistics
Filamentous actin
03 medical and health sciences
Stress
Physiological
Heat shock protein
Animals
Protein Interactions
Actin
030304 developmental biology
lcsh:R
Proteins
Actins
Rats
chemistry
lcsh:Q
Gene expression
030217 neurology & neurosurgery
Heat-Shock Response
Mathematics
Zdroj: PLoS ONE
PLoS ONE, Vol 8, Iss 7, p e68978 (2013)
ISSN: 1932-6203
Popis: Previous studies have suggested that the small heat shock protein, HspB1, has a direct influence on the dynamics of cytoskeletal elements, in particular, filamentous actin (F-actin) polymerization. In this study we have assessed the influence of HspB1 phosphorylation on its interaction(s) with F-actin. We first determined the distribution of endogenous non-phosphorylated HspB1, phosphorylated HspB1 and F-actin in neuroendocrine PC12 cells by immunocytochemistry and confocal microscopy. We then investigated a potential direct interaction between HspB1 with F-actin by precipitating F-actin directly with biotinylated phalloidin followed by Western analyses; the reverse immunoprecipitation of HspB1 was also carried out. The phosphorylation influence of HspB1 in this interaction was investigated by using pharmacologic inhibition of p38 MAPK. In control cells, HspB1 interacts with F-actin as a predominantly non-phosphorylated protein, but subsequent to stress there is a redistribution of HspB1 to the cytoskeletal fraction and a significantly increased association of pHspB1 with F-actin. Our data demonstrate HspB1 is found in a complex with F-actin both in phosphorylated and non-phosphorylated forms, with an increased association of pHspB1 with F-actin after heat stress. Overall, our study combines both cellular and biochemical approaches to show cellular localization and direct demonstration of an interaction between endogenous HspB1 and F-actin using methodolgy that specifically isolates F-actin.
Databáze: OpenAIRE
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