Biomedical subjects

David C Wong-Faull

Publications and source records attributed to David C Wong-Faull.

2 recordsLinked to original sources

PubMed2006

Nerve growth factor regulates adrenergic expression.

The mechanism by which nerve growth factor (NGF) regulates adrenergic expression was examined in PC-12 cells transfected with a rat phenylethanolamine N-methyl-transferase (PNMT) promoter-luciferase reporter gene construct pGL3RP893. NGF treatment increased PNMT promoter-driven luciferase activity in a dose- and time-dependent manner. Induction was attenuated by inhibition of the extracellular signal-regulated kinase mitogen-activated protein kinase (MAPK) pathway ( approximately 60%) but not by inhibition of the protein kinase A (PKA), protein kinase C, phosphoinositol kinase, or p38 MAPK pathways. Deletion PNMT promoter-luciferase reporter gene constructs showed that the NGF-responsive sequences lay within the proximal -392 base pairs (bp) of PNMT promoter, wherein binding elements for Egr-1 (-165 bp) and Sp1 (-48 bp) reside. Western analysis further showed that NGF increased nuclear levels of Egr-1, but not Sp1 or the catalytic subunit of PKA. Gel mobility shift assays showed increased potential for Egr-1, but not Sp1, protein-DNA binding complex formation. Mutation of either the Egr-1 or Sp1 binding sites in the PNMT promoter attenuated NGF activation. NGF, combined with pituitary adenylyl cyclase-activating protein (PACAP), another PNMT transcriptional activator, cooperatively stimulated PNMT promoter driven-luciferase activity beyond levels observed with either neurotrophin alone. Finally, post-transcriptional control seems to be another important mechanism by which neurotrophins regulate the adrenergic phenotype. NGF, PACAP, and a combination of the two stimulated both intron-retaining and intronless PNMT mRNA and PNMT protein, but to different extents.

Adrenal Glands↗

PubMed2004

Genetic mechanisms for adrenergic control during stress.

Cortisol and epinephrine released in response to stress are replenished via activation of the hypothalamic-pituitary-adrenal (HPA or stress) axis. Immobilization (IMMO) stress in rats stimulates epinephrine production in part via the gene encoding the epinephrine-synthesizing enzyme phenylethanolamine N-methyltransferase (PNMT). PNMT mRNA rose up to 7.0-fold with acute or chronic stress. Two transcription factors mediating stress induction of the PNMT gene are the glucocorticoid receptor (GR) and Egr-1, which interact with -533, -759, and -773 bp, and -165 bp binding sites in the rat PNMT promoter, respectively. To identify molecular mechanisms involved, effects of hypoxic stress on PNMT promoter activity were examined in PC12 cells transfected with the PNMT promoter-luciferase reporter gene construct pGL3RP893. Oxygen reduction to 5% increased PNMT promoter-driven luciferase expression, with maximum activity at 6 h. Pretreatment of the cells with protein kinase A (PKA) and protein kinase C (PKC) inhibitors, H-89 and GF109203X, respectively, attenuated the rise in luciferase. Similarly, PKA-deficient PC12 cells transfected with pGL3RP893 and exposed to hypoxia also showed attenuated PNMT promoter-driven luciferase expression. Mutation of the Egr-1 binding site completely prevented PNMT promoter activation, indicating that Egr-1 is essential to the stress response. Consistent with this result, hypoxia increased Egr-1 protein. Hypoxia also increased endogenous PNMT mRNA. However, a shift to intron-retaining mRNA from which truncated, nonfunctional protein is produced, occurred, suggesting that posttranscriptional regulation may be an important genetic mechanism controlling adrenergic expression and hence, epinephrine, during stress.

Animals↗