NERVE GROWTH-FACTOR REGULATION OF M4 MUSCARINIC RECEPTOR MESSENGER-RNA STABILITY BUT NOT GENE-TRANSCRIPTION REQUIRES MITOGEN-ACTIVATED PROTEIN-KINASE ACTIVITY

Nerve growth factor (NGF) up-regulated steady-state levels of m4 musca rinic acetylcholine receptor (mAChR) mRNA in PC12 cells. Up-regulation of mRNA levels was associated with a corresponding increase in mAChR binding sites. Two other growth factors, basic fibroblast growth facto r (bFGP) and epidermal growth factor (EGF), up regulated m4 mRNA and m AChR binding sites. Treatment of PC12 cells with NGF and bFGF, but not EGF, has previously been demonstrated to result in sustained activati on of mitogen activated protein kinase (MAPK), Analogously, NGF and bF GF, but not EGF, increased the stability of m4 mRNA in PC12 cells. In HER-PC12 cells, a clonal PC12 cell transfectant overexpressing EGF rec eptors and displaying sustained MAPK activation upon receptor stimulat ion, EGF treatment stabilized the m4 transcript, A synthetic inhibitor of MAPK kinase, PD98059, inhibited growth factor-induced stabilizatio n of the m4 transcript in both PC12 and HER-PC12 cells. These findings demonstrate that the MAPK pathway is involved in transcript stabiliza tion. Cycloheximide pretreatment abolished the posttranscriptional eff ect of NGF, indicating that de novo protein synthesis was required for the observed increase in m4 mRNA stability, By contrast, cycloheximid e had no discernible post-transcriptional effect if added after NGF tr eatment, suggesting that an inducible yet stable protein factor was in volved in m4 mRNA decay, An unusually well conserved 137 nucleotides o f m4 3'-untranslated region has been identified by sequence comparison with other mRNAs that are post-transcriptionally regulated by NGF. In PC12 cells that heterologously overexpress this region, we demonstrat e that NGF no longer stabilizes endogenous m4 mRNA This conserved regi on probably represents an NGF-responsive element involved in mRNA stab ility regulation, Finally, transcription of the m4 gene can be induced by all three growth factors but is not dependent on MAPK activity, un like growth factor-induced m4 mRNA stabilization.