Abstract
Abstract: :
Purpose: Alpha crystallins are distinct anti-apoptotic regulators as demonstrated by several laboratories. Regarding the mechanism, our recent work demonstrated that alphaB-crystallin represses caspase-3 activation during okadaic acid-induced apoptosis (Li et al., 2001. Exp. Cell Res. 266, 279-291). Furthermore, we found that alphaB-crystallin is able to interact with procaspase-3 and partially processed procaspase-3 (Mao et al., 2001, J. Biol Chem. 276, 43435-43445). Here we have demonstrated that during calcimycin-induced apoptosis, alphaB-crystallin is able to repress activities of ERK/p38 kinases and modify expression of several genes. Methods: The parental, vector- and alphaB-crystallin-transfected N/N1003A cells were treated with calcimycin for different length of time. The relative percentage of apoptotic cells was determined with viability assay. The total proteins and activity levels of ERK/p38 kinases in 3 different types of cells were examined with Western blot analysis. Expression of Bcl-Xs and other genes was also examined with Western blot analysis. Results: During treatment by calcimycin from 1 to 5 hours, ERK1/2 and p38 were stably activated in both N/N1003A and vector-transfected cells. However, in mouse alphaB-transfected cells, the activities of both ERK/p38 MAP kinases were significantly down-regulated after one-hour treatment and these low levels of activities were maintained in the next few hours. Associated with differential ERK/p38 activities in 3 different types of cells, we observed differential cell death rate. For example, after 5-hour treatment, only half of the apoptotic cells was found in alphaB-transfected cells than in parental or vector-transfected N/N1003A cells. Bcl-Xs was greatly up-regulated in parental or vector-transfected cells. In contrast, in alphaB-crystallin expression cells, Bcl-Xs expression displayed little change. Conclusion: AlphaB-crystallin-induced repression of ERK/JNK MAP kinase activity is part of the mechanism for prevention of calcimycin-induced apoptosis. Supported by EY 11372, NJ Foundation and UMDNJ/GSBS. None.
Keywords: 378 crystallins • 341 cell death/apoptosis • 580 signal transduction