Impaired nuclear accumulation and shortened phosphorylation of ERK after growth factor stimulation in cultured hepatocytes from rats exposed to 2-acetylaminofluorene

The hepatic carcinogen 2-acetylaminofluorene (AAF) exerts its effect as a t umor promoter by mitoinhibition of normal hepatocytes. Initiated cells prol iferate selectively and develop into preneoplastic foci and subsequently in to carcinomas. To study whether some of the mitoinhibitory effects of AAF c ould be attributed to an influence on intracellular signal transduction, gr owth factor signaling was studied in cultured hepatocytes from rats fed AAF for 7 d. Activation through the epidermal growth factor receptor (EGFR) wa s used to probe possible changes in downstream mitogenic signaling mechanis ms. The proliferative response to epidermal growth factor (EGF), measured a s proliferating cell nuclear antigen expression and thymidine incorporation , was almost completely inhibited in hepatocytes exposed to AAF. Neither EG FR protein revels nor EGF binding was notably altered in AAF-exposed hepato cytes as opposed to normal hepatocytes. The initial tyrosine phosphorylatio n of EGFR and downstream activation of Sos, Raf-1, and extracellular signal -regulated protein kinase (ERK) were similar in AAF-treated and control hep atocytes. Even though ERK phosphorylation was unaffected, a remarkable (80% ) reduction of ERK nuclear accumulation was observed in AAF-exposed hepatoc ytes immediately after mitogen stimulation. EGFR tyrosine phosphorylation a nd downstream signaling lasted 6 h in control cells Versus 2 h in AAF-expos ed hepatocytes. We previously demonstrated that AAF inhibits the growth fac tor-dependent induction of cyclin D1 and arrests hepatocyte cell-cycle prog ression before the p21/CIP1-controlled DNA-damage check point. The present data indicate that the DNA-damaging carcinogen AAF induces growth inhibitio n by a distinct inhibition of ERK nuclear accumulation after mitogen stimul ation. Inhibition of intracellular signal transduction may represent a nove l mechanism of growth arrest. Mel. Carcinog. 28:84-96, 2000. (C) 2000 Wiley -Liss, Inc.