Section 35
Chapter 34,967

GCM regulates the phenylethanolamine N-methyltransferase gene

Wong, D.L.; Tai, T.C.

FASEB Journal 18(4-5): Abstract 174


ISSN/ISBN: 0892-6638
Accession: 034966682

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To determine molecular mechanisms by which stress controls epinephrine and PNMT, we have been identifying transcriptional regulators of the PNMT gene critical to its stress response. A glial cell missing factor (GCM) has been linked to PNMT gene induction via protein kinase A (PKA) signaling. Gel mobility shift assays identified a PKA-sensitive response element at -440 bp in the rat PNMT promoter with a 3 bp mismatch to the 11 bp GCM binding element. In transfection assays with PNMT promoter-luciferase reporter gene constructs and GCM expression constructs in PC12 cells, GCM activated PNMT promoter-driven transcription. Deletion constructs identified a major activation site between -442 to -893 bp and a weaker site between -60 to -392 bp. Forskolin, but not PMA, stimulated PNMT promoter activity beyond levels seen with GCM alone and the combination only activated the promoter to the extent of forskolin. Mutation of the upstream GCM site markedly attenuated PNMT promoter activity to a greater extent than the forskolin-induced rise, although the ratio of PNMT promoter driven-luciferase expression in forskolin-stimulated to control cells remained constant. However, western analysis with GCM antibody did not detect GCM protein in PC12 cells despite GCM binding complex formation. Thus, a GCM-like factor may be responsible for PNMT promoter activation, rather than GCM itself.

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