Nerve Growth Factor, But Not Epidermal Growth Factor, Increases Fra-2 Expression and Alters Fra-2/JunD Binding to AP-1 and CREB Binding Elements in Pheochromocytoma (PC12) Cells

Abstract

In pheochromocytoma (PC12) cells nerve growth factor (NGF) and epidermal growth factor (EGF) activate similar receptor tyrosine kinase signaling pathways but evoke strikingly different biological outcomes: NGF induces differentiation and EGF acts as a mitogen. A novel approach was developed for identifying transcription factor activities associated with NGF-activated, but not EGF-activated, signaling, using random oligonucleotide clones from a DNA recognition library to isolate specific DNA binding proteins from PC12 nuclear extracts. A protein complex from NGF-treated, but not EGF-treated, cells was identified that exhibits increased mobility and DNA binding activity in gel mobility shift assays. The binding complex was identified in supershift assays as Fra-2/JunD. The clones used as probes contain either AP-1 or cAMP response element binding (CREB) recognition elements. Time course experiments revealed further differences in NGF and EGF signaling in PC12 cells. NGF elicits a more delayed and sustained ERK phosphorylation than EGF, consistent with previous reports. Both growth factors transiently induce c-fos, but NGF evokes a greater response than EGF. NGF specifically increases Fra-1 and Fra-2 levels at 4 and 24 hr. The latter is represented in Western blots by bands in the 40-46 kDa range. NGF, but not EGF, enhances the upper bands, corresponding to phosphorylated Fra-2. These findings suggest that prolonged alterations in Fra-2 and subsequent increases in Fra-2/JunD binding to AP-1 and CREB response elements common among many gene promoters could serve to trigger broadly an NGF-specific program of gene expression.

Chronic Nicotine Treatment Leads to Sustained Stimulation of Tyrosine Hydroxylase Gene Transcription Rate in Rat Adrenal Medulla

“…at ASPET Journals on May 7, 2018 jpet.aspetjournals.org NGF was shown to induce Fra-2 in PC12 cells (Boss et al, 2001). When rats were treated chronically with either saline or nicotine for 7 days and then administered a challenge injection of nicotine in the morning of the 8th day, the densities of the slowest migrating, Fra-2 immunoreactive bands increased dramatically 2 h after the challenge nicotine injection (S/N2 or N/N2 in Fig.…”

AP-1 proteins in the adult brain: facts and fiction about effectors of neuroprotection and neurodegeneration

“…Dierential eects of Jun proteins have been described for the regulation of hippocampal NGF expression following limbic seizures with participation of JunD in activation and of JunB in suppression (Elliott and Gall, 2000), and JunD might exert a major role in NGF production with variable AP-1 partners such as c-Fos or phosphorylated Fra-2 (Boss et al, 2001).…”

Alterations in Fos-Related Antigen 2 and σ1 Receptor Gene and Protein Expression Are Associated with the Development of Cocaine-Induced Behavioral Sensitization: Time Course and Regional Distribution Studies

“…Similar to other members of the fos family, Fra-2 is known to form heterodimeric complexes with other proteins, such as Jun and CREB family members (Herdegen and Leah, 1998;Smith et al, 2001). Moreover, Fra-2 can regulate the expression of other genes through interactions with the AP-1 DNA consensus element (Foletta, 1996;Boss et al, 2001). Thus, it is significant that the promoter of the 1 receptor contains putative AP-1 binding sites (Seth et al, 1997) through which Fra-2 could theoretically stimulate transcription.…”