DNA-dependent protein kinase is involved in heat shock protein-mediated accumulation of hypoxia-inducible factor-1alpha in hypoxic preconditioned HepG2 cells.
Hypoxic preconditioning may afford protection against subsequent lethal hypoxia. As hypoxic tolerance induces changes in the expression of genes involved in DNA damage and repair response pathways, we investigated whether DNA-dependent protein kinase (DNA-PK), one of the DNA double-strand break repair proteins, could be involved in hypoxic preconditioning-induced protective signaling cascades. We showed that induction of hypoxia-inducible factor-1alpha expression during hypoxic preconditioning by repeated hypoxic exposure was associated with increased mRNA and protein levels of DNA-PK catalytic subunit (DNA-PKcs) and Ku70/Ku80, the DNA-PK components, in human hepatoma HepG2 cells, followed by upregulation of Hsp70/Hsp90 and Bcl-2 and concurrent downregulation of Bax. Additionally, loss of DNA-PKcs led to attenuated expression of Hsp70/Hsp90, accelerated hypoxia-inducible factor-1alpha degradation, and increased susceptibility to hypoxia-induced cell death. We also found that the mRNA and protein levels of heat shock factor-1 (HSF1) were progressively increased with DNA-PK activation during hypoxic preconditioning, and inhibition of HSF1 function by KNK437 resulted in a significant decrease in the level of protein kinase Akt as well as of DNA-PKcs, with downregulation of Hsp70/Hsp90 and HIF-1alpha. Our results suggest the possibility that DNA-PK-mediated signaling pathway is required for the increase in HIF-1alpha expression through activation of HSF1 and subsequent upregulation of heat shock proteins after hypoxic reconditioning.
Kang
Moon Jung
MJ
Department of Biochemistry, Pusan National University School of Medicine, Busan, South Korea.
Jung
Sun Min
SM
Kim
Mi Ju
MJ
Bae
Jae Ho
JH
Kim
Hak Bong
HB
Kim
Joo Young
JY
Park
Soo Jung
SJ
Song
Hye Soon
HS
Kim
Dong Wan
DW
Kang
Chi Dug
CD
Kim
Sun Hee
SH
D000818:Animals; D034961:Antigens, Nuclear / Q000235:genetics / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics / Q000502:physiology; D001559:Benzhydryl Compounds / Q000494:pharmacology; D015153:Blotting, Western; D015687:Cell Hypoxia / Q000502:physiology; D002461:Cell Line, Transformed; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D051747:DNA-Activated Protein Kinase / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism; D000076249:Heat Shock Transcription Factors; D006360:Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics / Q000378:metabolism*; D000072200:Ku Autoantigen; D051379:Mice; D018345:Mice, Knockout; D016513:Mice, SCID; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D011760:Pyrrolidinones / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D051028:bcl-2-Associated X Protein / Q000235:genetics / Q000378:metabolism
D034961:Antigens, Nuclear; D001559:Benzhydryl Compounds; D015853:Cysteine Proteinase Inhibitors; D004268:DNA-Binding Proteins; C497442:HIF1A protein, human; C000619706:HSF1 protein, human; D018840:HSP70 Heat-Shock Proteins; D018841:HSP90 Heat-Shock Proteins; D000076249:Heat Shock Transcription Factors; D006360:Heat-Shock Proteins; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; C410027:KNK 437; D019253:Proto-Oncogene Proteins c-bcl-2; D011760:Pyrrolidinones; D014157:Transcription Factors; D051028:bcl-2-Associated X Protein; D051747:DNA-Activated Protein Kinase; C000606509:Xrcc6 protein, human; C000606510:Xrcc6 protein, mouse; D000072200:Ku Autoantigen