The cAMP responsive element and CREB partially mediate the response of the tyrosine hydroxylase gene to phorbol ester

Tyrosine hydroxylase (TH) gene promoter activity is increased in PC12 cells that are treated with the phorbol ester, 12-O-tetradecanoylphorbol 13-acetate (TPA). Mutagenesis of either the cAMP responsive element (CRE) or the activator protein-1 element (AP1) within the TH gene proximal promoter leads to a dramatic inhibition of the TPA response. The TH CRE and TH AP1 sites are also independently responsive to TPA in minimal promoter constructs. TPA treatment results in phosphorylation of cAMP responsive element binding protein (CREB) and activation of cAMP-dependent protein kinase (PKA) in PC12 cells; hence, we tested whether CREB and/or PKA are essential for the TPA response. In CREB-deficient cells, the response of the full TH gene proximal promoter or the independent response of the TH CRE by itself to TPA is inhibited. The TPA-inducibility of TH mRNA is also blocked in CREB-deficient cells. Expression of the PKA inhibitor protein, PKI, also inhibits the independent response of the TH CRE to TPA. Our results support the hypothesis that TPA stimulates the TH gene promoter via signaling pathways that activate either the TH AP1 or TH CRE sites. Both signaling pathways are dependent on CREB and the TH CRE-mediated pathway is dependent on PKA.     

The expression pattern of CREB genes in the central nervous system of the pond snail Lymnaea stagnalis

To analyze the expression pattern of genes of cAMP responsive element binding protein (CREB), we performed in situ hybridization for the whole central nervous system (CNS) of the pond snail Lymnaea stagnalis. The CREB1 (activator) and CREB2 (repressor) homologues have already been cloned in L. stagnalis, and they are referred to as LymCREB1 and LymCREB2. Using the frozen sections and the whole mount preparations of the CNS, we mapped the distribution of LymCREB1 and LymCREB2 mRNA containing neurons. The present findings showed that the LymCREB1 mRNA containing neurons are a relatively few, whereas LymCREB2 mRNA is contained ubiquitously in the whole CNS of L. stagnalis.     

Circadian expression of steroidogenic cytochromes P450 in the mouse adrenal gland--involvement of cAMP-responsive element modulator in epigenetic regulation of Cyp17a1

The cytochrome P450 (CYP) genes Cyp51, Cyp11a1, Cyp17a1, Cyb11b1, Cyp11b2 and Cyp21a1 are involved in the adrenal production of corticosteroids, whose circulating levels are circadian. cAMP signaling plays an important role in adrenal steroidogenesis. By using cAMP responsive element modulator (Crem) knockout mice, we show that CREM isoforms contribute to circadian expression of steroidogenic CYPs in the mouse adrenal gland. Most striking was the CREM-dependent hypomethylation of the Cyp17a1 promoter at zeitgeber time 12, which resulted in higher Cyp17a1 mRNA and protein expression in the knockout adrenal glands. The data indicate that products of the Crem gene control the epigenetic repression of Cyp17 in mouse adrenal glands.     

Regulation of gene expression in PC12 cells via an activator of dual second messengers: pituitary adenylate cyclase activating polypeptide.

In this study we demonstrate that the activator protein-1 (AP-1) DNA motif, initially considered to be unresponsive to cyclic AMP (cAMP), does function as a cAMP-response element in PC12 cells. A luciferase reporter gene driven by the collagenase promoter that contains the AP-1 motif is responsive to cAMP as well as phorbol esters when transfected in PC12 cells. We have recently shown that pituitary adenylate cyclase activating peptide (PACAP) has neurotrophic properties and activates both adenylylcyclase and the inositol lipid cascade in PC12 cells. Consistent with these actions, we demonstrate that PACAP is an effective activator of luciferase reporter genes whose promoters bear the AP-1 motif, as well as the related DNA element that binds the protein CREB. Both the cAMP and inositol lipid pathways appear to play a role in the activation of these motifs by PACAP. Mutation of the AP-1 motif and its juxtaposition to a heterologous promoter proves that the AP-1 motif is a locus for response to cAMP and PACAP. The luciferase reporter genes bearing the AP-1 motif are not cAMP responsive in HeLa tk- cells, indicating that the mode of second-messenger responsiveness is cell-type specific.