Site-directed mutagenesis of the ATM promoter: Consequences for response to proliferation and ionizing radiation
Gueven, N and Keating, K and Fukao, T and Loeffler, H and Kondo, N and Rodemann, HP and Lavin, MF, Site-directed mutagenesis of the ATM promoter: Consequences for response to proliferation and ionizing radiation, Genes, Chromosomes and Cancer, 38, (2) pp. 157-167. ISSN 1045-2257 (2003) [Refereed Article]
Although ATM, the protein defective in ataxia-telangiectasia (A-T), is activated primarily by radiation, there is also evidence that expression of the protein can be regulated by both radiation and growth factors. Computer analysis of the ATM promoter proximal 700-bp sequence reveals a number of potentially important cis-regulatory sequences. Using nucleotide substitutions to delete putative functional elements in the promoter of ATM, we examined the importance of some of these sites for both the basal and the radiation-induced activity of the promoter. In lymphoblastoid cells, most of the mutations in transcription factor consensus sequences [Sp1(1), Sp1(2), Cre, Ets, Xre, γlre(2), a modified AP1 site (Fse), and GCF] reduced basal activity to various extents, whereas others [γlre(1), NF1, Myb] left basal activity unaffected. In human skin fibroblasts, results were generally the same, but the basal activity varied up to 8-fold in these and other cell lines. Radiation activated the promoter approximately 2.5-fold in serum-starved lymphoblastoid cells, reaching a maximum by 3 hr, and all mutated elements equally blocked this activation. Reduction in Sp1 and AP1 DNA binding activity by serum starvation was rapidly reversed by exposure of cells to radiation. This reduction was not evident in A-T cells, and the response to radiation was less marked. Data provided for interaction between ATM and Sp1 by protein binding and co-immunoprecipitation could explain the altered regulation of Sp1 in A-T cells. The data described here provide additional evidence that basal and radiation-induced regulation of the ATM promoter is under multifactorial control.