Activation of mitochondrial promoter P(H)-binding protein in a radio-resistant Chinese hamster cell strain associated with Bcl-2.
The cellular response to ionizing radiation is mediated by a complex interaction of number of proteins involving different pathways. Previously, we have shown that up regulation of mitochondrial genes ND1, ND4, and COX1 transcribed from the heavy strand promoter (P(H)) has been increased in a radio-resistant cell strain designated as M5 in comparison with the parental Chinese hamster V79 cells. These genes are also up regulated in Chinese hamster V79 cells VB13 that express exogenous human Bcl2. In the present study, the expression of the gene ND6 that is expressed from the light strand promoter (P(L)) was found to be similar in both the cell lines, as determined by RT-PCR. To test the possibility that this differential expression of mitochondrial genes under these two promoters was mediated by differences in proteins' affinity to interact with these promoters, we have carried out electrophoretic mobility shift assay (EMSA) using mitochondrial cell extracts from these two cell lines. Our result of these experiments revealed that two different proteins formed complex with the synthetic promoters and higher amount of protein from M5 cell extracts interacted with the P(H) promoter in comparison to that observed with cell extracts from Chinese hamster V79 cells. The promoter-specific differential binding of proteins was also observed in VB13. These results showed that differential mitochondrial gene expression observed earlier in the radio-resistant M5 cells was due to enhanced interaction proteins with the promoters P(H) and mediated by the expression of Bcl2.
Roychoudhury
Paromita
P
Molecular and Human Genetics Division, Indian Institute of Chemical Biology, 4 Raja S.C. Mullick Road, Kolkata 700032, India.
Ghosh
Utpal
U
Bhattacharyya
Nitai P
NP
Chaudhuri
Keya
K
D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D004268:DNA-Binding Proteins / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D005720:Gamma Rays; D015971:Gene Expression Regulation, Enzymologic; D050259:Genes, Mitochondrial; D006801:Humans; D008297:Male; D008928:Mitochondria / Q000201:enzymology / Q000528:radiation effects; D024101:Mitochondrial Proteins / Q000378:metabolism*; D009245:NADH Dehydrogenase / Q000096:biosynthesis / Q000235:genetics; D011401:Promoter Regions, Genetic; D021122:Protein Subunits / Q000096:biosynthesis / Q000235:genetics; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D011836:Radiation Tolerance
D004268:DNA-Binding Proteins; D024101:Mitochondrial Proteins; D021122:Protein Subunits; D019253:Proto-Oncogene Proteins c-bcl-2; D009245:NADH Dehydrogenase