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Lesion-induced increase in nerve growth factor mRNA is mediated by c-fos
B Hengerer1, D Lindholm, R Heumann
1Max Planck Institute for Psychiatry, Department of Neurochemistry, Martinsried/Munich, Federal Republic of Germany.
Abstract:
Lesion of the sciatic nerve caused a rapid increase in c-fos and c-jun mRNA that was followed about 2 hr later by an increase in nerve growth factor (NGF) mRNA. To evaluate whether the initial increase in c-fos mRNA is causally related to the subsequent increase in NGF mRNA, we performed experiments with fibroblasts of transgenic mice carrying an exogenous c-fos gene under the control of a metallothionein promoter. In primary cultures of these fibroblasts, CdCl2 evoked a rapid increase in exogenous c-fos mRNA, followed immediately by an increase in endogenous c-jun mRNA and with a slight delay by an increase in NGF mRNA. In fibroblasts of C3H control mice, CdCl2 had no effect on the mRNA levels of the protooncogenes c-fos and c-jun or of NGF. Additional evidence for a causal relationship between c-fos induction and the subsequent increase in NGF mRNA was obtained in cotransfection experiments. Fibroblasts of C3H control mice were cotransfected with a metallothionein-promoter-driven c-fos expression vector and a NGF promoter-chloramphenicol acetyltransferase reporter gene construct. Induction of the exogenous c-fos by CdCl2 resulted in increased activity of the NGF promoter. DNase I footprint experiments demonstrated that a binding site for transcription factor AP-1 (Fos/Jun heterodimer) in the first intron of the NGF gene was protected following c-fos induction. That this protected AP-1 site indeed was functional in the regulation of NGF expression was verified by deletion experiments and by a point mutation in the corresponding AP-1 binding region in the NGF promoter-chloramphenicol acetyltransferase reporter construct.
Insights
Sciatic nerve injury rapidly increases c-fos and c-jun mRNA, leading to nerve growth factor (NGF) mRNA increase. This study confirms c-fos directly upregulates NGF mRNA via AP-1 transcription factor binding.
Area of Science:
- Neurobiology
- Molecular Biology
- Gene Regulation
Background:
- Sciatic nerve injury triggers rapid expression of immediate-early genes like c-fos and c-jun.
- Nerve growth factor (NGF) mRNA levels increase following nerve injury, but the regulatory mechanisms are not fully understood.
Purpose of the Study:
- To investigate the causal relationship between c-fos gene induction and the subsequent upregulation of nerve growth factor (NGF) mRNA.
- To elucidate the role of the transcription factor AP-1 in mediating c-fos-induced NGF expression.
Main Methods:
- Experiments using fibroblasts from transgenic mice with an inducible c-fos gene.
- Cadmium chloride (CdCl2) induction of c-fos expression and measurement of c-fos, c-jun, and NGF mRNA levels.
- Cotransfection assays with NGF promoter-reporter constructs and DNase I footprinting to identify transcription factor binding sites.
Main Results:
- CdCl2 induced exogenous c-fos mRNA, followed by increased endogenous c-jun and NGF mRNA in transgenic fibroblasts.
- Control fibroblasts showed no change in mRNA levels upon CdCl2 treatment.
- Induction of exogenous c-fos in control fibroblasts enhanced NGF promoter activity, and DNase I footprinting identified a functional AP-1 binding site in the NGF gene intron.
Conclusions:
- c-fos induction is causally linked to the subsequent increase in NGF mRNA expression.
- The transcription factor AP-1 plays a critical role in mediating c-fos-dependent regulation of NGF.
- This study reveals a novel molecular mechanism for NGF gene regulation following nerve injury.