α-Crystallin B prevents apoptosis after H2O2 exposure in mouse neonatal cardiomyocytes.
α-Crystallin B (cryAB) is the most abundant small heat shock protein in cardiomyocytes (CMs) and has been shown to have potent antiapoptotic properties. Because the mechanism by which cryAB prevents apoptosis has not been fully characterized, we examined its protective effects at the cellular level by silencing cryAB in mouse neonatal CMs using lentivector-mediated transduction of short hairpin RNAs. Subcellular fractionation of whole hearts showed that cryAB is cytosolic under control conditions, and after H(2)O(2) exposure, it translocates to the mitochondria. Phosphorylated cryAB (PcryAB) is mainly associated with the mitochondria, and any residual cytosolic PcryAB translocates to the mitochondria after H(2)O(2) exposure. H(2)O(2) exposure caused increases in cryAB and PcryAB levels, and cryAB silencing resulted in increased levels of apoptosis after exposure to H(2)O(2). Coimmunoprecipitation assays revealed an apparent interaction of both cryAB and PcryAB with mitochondrial voltage-dependent anion channels (VDAC), translocase of outer mitochondrial membranes 20 kDa (TOM 20), caspase 3, and caspase 12 in mouse cardiac tissue. Our results are consistent with the conclusion that the cardioprotective effects of cryAB are mediated by its translocation from the cytosol to the mitochondria under conditions of oxidative stress and that cryAB interactions with VDAC, TOM 20, caspase 3, and caspase 12 may be part of its protective mechanism.
Department of Physiology, University of Toronto, Toronto, Ontario, Canada M5G 1L6.
Roxana
R
Chis
Parveen
P
Sharma
Nicolas
N
Bousette
Tetsuaki
T
Miyake
Aaron
A
Wilson
Peter H
PH
Backx
Anthony O
AO
Gramolini
D000367:Age Factors; D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D006861:Hydrogen Peroxide / Q000633:toxicity*; D051379:Mice; D008928:Mitochondria / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D016877:Oxidants / Q000633:toxicity; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology*; D010766:Phosphorylation / Q000502:physiology; D034741:RNA, Small Interfering / Q000235:genetics; D017382:Reactive Oxygen Species / Q000378:metabolism; D038203:alpha-Crystallin B Chain / Q000235:genetics* / Q000378:metabolism
D016877:Oxidants; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; D038203:alpha-Crystallin B Chain; D006861:Hydrogen Peroxide