REGULATION OF EPIDERMAL GROWTH-FACTOR RECEPTOR BY ACTIVATED H-RAS ANDV-MYC ONCOGENES IN MOUSE BALB 3T3 CELLS - POSSIBLE ROLES OF AP-1/

We previously reported that introduction of H-ras oncogene decreases t he epidermal growth factor (EGF) binding activity to cell surface EGF receptor in mouse Balb/3T3, In this study, we have further isolated fo ur H-ras transfectants, four v-myc transfectants and three both H-ras and v-myc (H-ras/v-myc) transfectants of mouse Balb/3T3 cells. In comp arison with introduction of v-myc alone or both H-ras and v-myc oncoge nes, introduction of H-ras alone resulted in a loss of [I-125]EGF bind ing activity to the cell surface EGF receptor, RT-PCR analysis also sh owed much lower levels of EGF receptor gene expression in H-ras transf ectants compared to that of parental untransformed cells (Balb-Neo1), v-myc and H-ras/v-myc transfectants, Our results demonstrated the acti vated binding of a transcription factor, Stat1 p84/p91, which directly interacts with EGF receptor, to c-sis-inducible element (SIE) in both v-myc and H-ras/v-myc transfectants, but not in H-ras transfectants, Among transcription factors which we have analysed, activator protein 1 (AP-1) but not SP-1 was modulated by H-ras. Gel shift assays demonst rated the mobility pattern of TPA-responsive element (TRE) binding com plex with AP-1 derived from H-ras transfectants migrated faster than t hose from Balb-Neo1, v-myc and H-ras/v-myc. Expression of c-Jun and Fr a-1 was increased more than threefold in H-ras transfectants compared with Balb-Neo1, v-myc and H-ras/v-myc transfectants, but that of c-Fos , Jun B and SP-1 was unchanged, Both transient and permanent expressio n of H-ras enhanced AP-1 activity in mouse cells, but further co-intro duction of dominant negative c-jun mutant encoding a transcriptionally inactive product inhibited the H-ras dependent AP-1 induction, Transf ection of the dominant negative c-jun mutant also restored down-regula tion of EGF binding by activated H-ras oncogene, Down-regulation of EG F receptor by activated H-ras and the possible involvement of a transc ription factor, AP-1, will be discussed.