Selective degradation of aggregate-prone CryAB mutants by HSPB1 is mediated by ubiquitin-proteasome pathways.
Disease-causing mutations of genes encoding small MW heat shock proteins (sHSPs) constitute a growing family of inherited myofibrillar disorders. In the present work, we found that three structurally-distinct CryAB mutants R120G, 450delA and 464delCT are mostly present in the detergent-insoluble fractions when overexpressed in H9c2 rat heart cells. We found that either over-expression or knockdown of HSPB1, a related sHSP, affects the solubility, stability, and degradation of aggregation-prone CryAB mutants. HSPB1 overexpression has negligible effects on the solubility and protein aggregates of either R120G and/or 450delA but increased the solubility and prevented formation of 464delCT aggregates. HSPB1 knockdown decreased solubility and increased protein aggregates of all CryAB mutants, indicating a key role for HSPB1 in clearance of CryAB mutants under basal conditions. We provide four lines of evidence that such selective clearance of R120G, 450delA and 464delCT mutants by HSPB1 is mediated by the ubiquitin-proteasome system (UPS). First, we found that treatment with the proteasome inhibitors increased the levels of all CryAB mutants. Second, R120G and 450delA overexpression corresponded to the accumulation of their specific ubiquitin conjugates in H9c2 cells. Third, HSPB1 knockdown directly increased the levels of all polyubiquitin conjugates. And fourth, the selective attenuation of 464delCT expression by HSPB1 over-expression was abrogated by the proteasome inhibition. We conclude that such selective interactions between CryAB mutants and HSPB1 overexpression might have important implications for the clinical manifestations and potential treatment.
Zhang
Huali
H
Center for Cardiovascular Translational Biomedicine, Division of Cardiology, Department of Internal Medicine, University of Utah School of Medicine, Salt Lake City, UT 84132, USA.
Rajasekaran
Namakkal S
NS
Orosz
Andras
A
Xiao
Xianzhong
X
Rechsteiner
Martin
M
Benjamin
Ivor J
IJ
D000595:Amino Acid Sequence; D000818:Animals; D001343:Autophagy; D003593:Cytoplasm / Q000378:metabolism; D015536:Down-Regulation; D055785:Gene Knockdown Techniques; D055551:HSP27 Heat-Shock Proteins / Q000378:metabolism*; D006360:Heat-Shock Proteins; D006801:Humans; D051379:Mice; D018832:Molecular Chaperones; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000166:cytology / Q000378:metabolism; D025821:Polyubiquitin / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D020836:Protein Structure, Quaternary; D051381:Rats; D015398:Signal Transduction; D012995:Solubility; D013347:Subcellular Fractions / Q000378:metabolism; D025801:Ubiquitin / Q000378:metabolism*; D038203:alpha-Crystallin B Chain / Q000737:chemistry* / Q000378:metabolism*
D055551:HSP27 Heat-Shock Proteins; C057725:HSPB1 protein, human; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D050505:Mutant Proteins; D025801:Ubiquitin; D038203:alpha-Crystallin B Chain; D025821:Polyubiquitin; D046988:Proteasome Endopeptidase Complex