IDENTIFICATION OF ACTIVE SUBSTANCES FROM STREPTOMYCES CULTURE FLUIDS USING P53-INDEPENDENT EXPRESSION OF P21 WAF1/CIP1 GENE AND THEIR MODE OF ACTION/

An assay system was constructed to identify chemicals that have a pote ntial to induce p21/WAF1 gene, a target of the tumor suppressor p53 cr itical for negative growth regulation, Screening of about 1300 culture fluids of Streptomyces resulted in identification of active substance s which induced the p21 gene in a p53-independent manner; one was a mi xture of four members of the actinomycin group, and the other was tric hostatin A, Transcriptional regulatory regions of p21 gene for inducti on by actinomycin D and trichostatin A were determined by transient ex pression of luciferase constructs in cells which are p53-deficient (Sa os-2) or express a mutated form of p53 (TMK-1). The essential transcri ptional elements for the response to these drugs localize within 210 b p of the 5'-upstream region of human p21 gene, and Sp1 elements were d etermined to be critical for the induction. DNA-binding activity of Sp 1 was not increased in cells treated with these drugs, but kinase inhi bitors such as staurosporin and wortmannin inhibited the induction.