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Papers In Press, published online ahead of print September 6, 2001
Molecular Biology, University of Medicine and Dentistry of New Jersey-SOM, Stratford, NJ 08084
Corresponding Author: DWL168{at}att.net
SUMMARY It is well established that the proto-oncogene, bcl-2, can prevent apoptosis induced by a variety of factors. Regarding the mechanism by which Bcl-2 prevents cell death, one theory suggests that it acts by protecting cells from oxidative stress. In the lens system, oxidative stress-induced apoptosis is implicated in cataractogenesis. To explore the possibility of anti-apoptotic gene therapy development for cataract prevention and also to further test the anti-oxidative stress theory of Bcl-2 action, we have introduced the human bcl-2 gene into an immortalized rabbit lens epithelial cell line, N/N1003A. The stable expression lines of both vector- and bcl-2-transfected cells have been established. Treatment of the two cell lines with H2O2 revealed that bcl-2-transfected cells were less capable of detoxifying H2O2 than the control cells. Moreover, bcl-2-transfected cells are more susceptible to H2O2-induced apoptosis. To explore why bcl-2-transfected cells have reduced resistance to H2O2-induced apoptosis, we examined the expression patterns of several relevant genes in both vector- and bcl-2-transfected cells and found that expression of the alphaB-crystallin gene was distinctly down-regulated in bcl-2-transfected cells. This down-regulation was specific since a substantial inhibition of Bcl-2 expression through antisense bcl-2 RNA significantly restored the level of aB-crystallin, and moreover, enhanced the ability of the bcl-2-transfected cells against H2O2-induced apoptosis. Introduction of a mouse aB-crystallin gene into bcl-2-transfected cells also counteracted the Bcl-2 effects. In Bcl-2 expression cells, the exogenous mouse aB-crystallin gene promoter linked to CAT reporter gene was also down-regulated. Down-regulation of aB-crystallin gene was largely derived from changed LEDGF activity. Besides, aB-crystallin prevents apoptosis through interaction with procaspase-3 and partially processed procaspase-3 to prevent caspase-3 activation. Together, our results reveal that Bcl-2 can regulate gene expression in rabbit lens epithelial cells. Through down-regulation of the aB-crystallin gene, Bcl-2 attenuates the ability of rabbit lens epithelial cells against H2O2-induced apoptosis.
J. Biol. Chem, 10.1074/jbc.M102195200
Submitted on March 12, 2001
Revised on September 6, 2001
Accepted on September 5, 2001
Human Bcl-2 gene attenuates the ability of rabbit lens epithelial cells against H2O2-induced apoptosis through down-regulation of the alphaB crystallin gene
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