THE MACROMOLECULAR STATE OF THE TRANSCRIPTION FACTOR E2F AND GLUCOCORTICOID REGULATION OF C-MYC TRANSCRIPTION

Glucocorticoids inhibit transcription of the proto-oncogene c-myc in l ymphoid cells of thymic origin. To determine if this effect is associa ted with changes in the properties of the transcription factor E2F, ex tracts were prepared from control and glucocorticoid-treated P1798 mur ine T lymphoma cells, and the macromolecular state of E2F was assessed by gel-mobility shift. Control extracts exhibit two predominant gel-m obility shift entities of which one corresponds to ''free'' E2F. A sec ond entity, complex C, has properties similar to those described for t he complex containing E2F, p107, cyclin A, and Cdk2. Complex C disappe ars after addition of dexamethasone and is replaced by complex D. The mobility of this complex and its sensitivity to SV40 T antigen suggest that complex D corresponds to an E2F.p105(Rb-1) complex. Extracts fro m control and glucocorticoid-treated cells yield identical DNase I pro tection patterns on the c-myc P2 promoter. Furthermore, such extracts transcribe the c-myc P2 promoter in vitro with equal activity. The rel ative abundance of the E2F complexes was measured after addition of de xamethasone. Complex C disappears as cells withdraw from S phase, and complex D appears at this time. The genes encoding thymidine kinase (T k-1) and p34(cdc2) (cdc2) are regulated with kinetics similar to those observed for changes in the macromolecular state of E2F. However, reg ulation of c-myc expression occurs long before any change in E2F. The macromolecular state of E2F may regulate expression of genes at the G( 1)/S boundary. However, the data are not consistent with the hypothesi s that association of E2F with tumor suppressor gene products such as p107 or p105(Rb-1) is relevant to glucocorticoid regulation of c-myc t ranscription.