Regulation of Bcl-xL expression by H2O2 in cardiac myocytes.
Oxidative stress promotes cardiac myocyte apoptosis through the mitochondrial death pathway. Since Bcl-2 family proteins are key regulators of apoptosis, we examined the effects of H2O2 on the expression of principal Bcl-2 family proteins (Bcl-2, Bcl-xL, Bax, Bad) in neonatal rat cardiac myocytes. Protein expression was assessed by immunoblotting. Bcl-2, Bax, and Bad were all down-regulated in myocytes exposed to 0.2 mm H2O2, a concentration that induces apoptosis. In contrast, although Bcl-xL levels initially declined, the protein was re-expressed from 4-6 h. Bcl-xL mRNA was up-regulated from 2 to 4 h in neonatal rat or mouse cardiac myocytes exposed to H2O2, consistent with the re-expression of protein. Four different untranslated first exons have been identified for the Bcl-x gene (exons 1, 1B, 1C, and 1D, where exon 1 is the most proximal and exon 1D the most distal to the coding region). All were detected in mouse or rat neonatal cardiac myocytes, but exon 1D was not expressed in adult mouse hearts. In neonatal mouse or rat cardiac myocytes, H2O2 induced the expression of exons 1B, 1C, and 1D, but not exon 1. These data demonstrate that the Bcl-x gene is selectively responsive to oxidative stress, and the response is mediated through distal promoter regions.
National Heart and Lung Institute Division (Cardiac Medicine Section), Faculty of Medicine, Imperial College London, Flowers Building, Armstrong Road, London SW7 2AZ, United Kingdom.
Donna M
DM
Valks
Timothy J
TJ
Kemp
Angela
A
Clerk
D000818:Animals; D017209:Apoptosis; D002478:Cells, Cultured; D017931:DNA Primers / Q000494:pharmacology; D018076:DNA, Complementary / Q000378:metabolism; D015536:Down-Regulation; D005091:Exons; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D015151:Immunoblotting; D051379:Mice; D008810:Mice, Inbred C57BL; D008957:Models, Genetic; D009206:Myocardium / Q000378:metabolism*; D018384:Oxidative Stress; D011401:Promoter Regions, Genetic; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis* / Q000235:genetics*; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013154:Spleen / Q000378:metabolism; D013997:Time Factors; D015854:Up-Regulation; D051020:bcl-X Protein
C494812:Bcl2l1 protein, mouse; C494813:Bcl2l1 protein, rat; D017931:DNA Primers; D018076:DNA, Complementary; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D051020:bcl-X Protein; D012313:RNA; D006861:Hydrogen Peroxide