NERVE GROWTH-FACTOR INDUCES TRANSCRIPTION OF NGFIA THROUGH COMPLEX REGULATORY ELEMENTS THAT ARE ALSO SENSITIVE TO SERUM AND PHORBOL 12-MYRISTATE 13-ACETATE

The expression of NGFIA (also known as egr1, zif268, TIS8, krox24, and d2) is rapidly and transiently increased by nerve growth factor (NGF) in PC12 cells. The 5'-region of this gene includes four serum response elements (SREs), a cAMP-like response element, an AP1-like response element, and an SP1-binding site. From deletion analysis of chloramphenicol acetyltransferase reporter constructs, we have established that the first 106 basepairs 5' of the transcriptional start site are sufficient for induction of NGFIA by NGF in PC12 cells; deletion beyond this point results in dramatically reduced induction of the gene. Using defined mutations in the NGFIA promoter and NGFIA-thymidine kinase hybrid promoters, we have defined three elements (SRE1, SRE2, and AP1-like) in the first 106 basepairs of upstream DNA, each of which contributes to induction of NGFIA by NGF. Cooperation by two of these elements (i.e. the two SREs or one SRE and the AP1-like element) is sufficient to confer transcriptional induction by NGF, but the combination of all three elements increased induction by NGF more effectively than a pair of elements. This suggests that the response of NGFIA to NGF is mediated by a cis-acting sequence that is composed of at least three distinct elements. An oligonucleotide composed of SRE1 and SRE2 that can confer the ability for NGF induction to heterologous promoter constructs complexes with proteins in PC12 cell nuclear extracts, but the protein-DNA complexes do not appear to be altered by NGF treatment, as measured by DNA mobility shift assays. We have also established that the regulatory region of NGFIA that mediates NGF induction also mediates the induction by serum and phorbol 12-myristate 13-acetate, suggesting that multiple signal transduction pathways must converge on these sequences to regulate the expression of this gene.