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The significance of phosphorylation in the stress‐induced translocation of αB‐crystallin
Author(s) -
Krishnamoorthy Vigneshwaran,
Donofrio Anthony John,
Martin Jody Lee
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.4.a385-b
Subject(s) - phosphorylation , mutant , chromosomal translocation , serine , cytosol , cytoskeleton , microbiology and biotechnology , myofilament , biology , heat shock protein , hsp70 , chemistry , biochemistry , myocyte , cell , enzyme , gene
αB‐crystallin is a member of the small heat shock protein family that is highly expressed in striated muscle tissues. Overexpression and knockout studies have suggested a cardioprotective role for αB‐crystallin. Stress‐induced translocation of αB‐crystallin to the cytoskeletal elements contributes to its cytoprotective role. In the heart, phosphorylation (at ser‐19, ser‐45 and ser‐59) and O‐GlcNAcylation (thr‐170) are the only two signaling events known so far. There are conflicting views on the significance of phosphorylation in triggering the αB‐crystallin translocation. To understand the role of phosphorylation, we have developed adenoviral vectors expressing non‐phosphorylatable (αB‐S19A, αB‐S45A, αB‐S59A and αB‐3A) and phosphomimicking mutants (αB‐2AE and αB‐3E) of αB‐crystallin. These mutants were transduced into neonatal rat ventricular myocytes, subjected to heat shock and lysed in HEPES buffered saline with 0.5% Triton X‐100. The detergent soluble (cytosol) and detergent insoluble (cytoskeleton) fractions were separated and immunoblotted for αB‐crystallin. The αB‐S59A mutant showed the greatest retention in the soluble fraction while the αB‐3E showed maximum translocation, even in the absence of stress. Similarly upon examining nuclear and post‐nuclear supernatants αB‐S45A mutant translocated to the nucleus much more robustly than other single alanine mutants. These data suggests that phosphorylation at each serine residue acts in an additive manner to facilitate stress‐induced translocation and also specifically targets αB‐crystallin. Funding: American Heart Association