The repertoire of Fos and Jun proteins expressed during the G(1) phase of the cell cycle is determined by the duration of mitogen-activated protein kinase activation

In Rat-1 fibroblasts nonmitogenic doses of lysophosphatidic acid (LPA) stim ulate a transient activation of mitogen-activated protein kinase (MAPK), wh ereas mitogenic doses elicit a sustained response. This sustained phase of MAPK activation regulates cell fate decisions such as proliferation or diff erentiation, presumably by inducing a program of gene expression which is n ot observed in response to transient MAPK activation. We have examined the expression of members of the AP-1 transcription factor complex in response to stimulation with different doses of LPA. c-Fos, c-Jun, and JunB are indu ced rapidly in response to LPA stimulation, whereas Fra-1 and Fra-2 are ind uced after a significant lag. The expression of c-Fos is transient, whereas the expression of c-Jun, JunB, Fra-1, and Fra-2 is sustained. The early ex pression of c-Fos can be reconstituted with nonmitogenic doses of LPA, but the response is transient compared to that observed with mitogenic doses. I n contrast, expression of Fra-1, Fra-2, and JunB and optimal expression of c-Jun are observed only with doses of LPA which induce sustained MAPK activ ation and DNA synthesis. LPA-stimulated expression of c-Fos, Fra-1, Fra-2, c-Jun, and JunB is inhibited by the MEK1 inhibitor PD098059, indicating tha t the Raf-MEK-MAPK cascade is required for their expression. In cells expre ssing a conditionally active form of Raf-l (Delta Raf-1:ER), we observed th at selective, sustained activation of Raf-MEK-MAPK was sufficient to induce expression of Fra-1, Fra-2, and JunB but, interestingly, induced little or no c-Fos or c-Jun. The induction of c-Fos observed in response to LPA was strongly inhibited by buffering the intracellular [Ca2+]. Moreover, althoug h Raf activation or calcium ionophores induced little c-Fos expression, we observed a synergistic induction in response to the combination of Delta Ra f-1:FR and ionomycin. These results suggest that kinetically distinct phase s of MAPK activation serve to regulate the expression of distinct AP-1 comp onents such that sustained MAPK activation is required for the induced expr ession of Fra-1, Fra-2, c-Jun, and JunB. However, in contrast to the case f or Fra-1, Fra-2, and JunB, activation of the MAPK cascade alone is not suff icient to induce c-Fos expression, which rather requires cooperation with o ther signals such as Ca2+ mobilization. Finally, the identification of the Fra-1, Fra-2, c-Jun, and JunB genes as genes which are selectively regulate d by sustained MAPK activation or in response to activated Raf suggests tha t they are candidates to mediate certain of the effects of Ras proteins in oncogenic transformation.