cAMP应答元件结合蛋白与痛觉调制

cAMP应答元件结合蛋白（cAMP response element binding protein,CREB）是刺激诱导的一种转录因子,通过磷酸化实现调节转录功能。疼痛和痛觉过敏是组织损伤或炎症时常伴有的生理病理过程,谷氨酸、P物质等神经递质或神经肽以及细胞内的信号转导途径参与此过程。近年来研究发现CREB通过自身磷酸化,在炎症、神经损伤等诱发的自发性疼痛、痛觉过敏及痛觉超敏中具有重要作用。本文从CREB的一般特性及其在脊髓水平的痛觉调制中的作用等方面予以综述。     

DNA replication stress-induced phosphorylation of cyclic AMP response element-binding protein mediated by ATM

The DNA damage-response regulators ATM (ataxia-telangiectasia-mutated) and ATR (ATM-Rad3-related) are structurally and functionally related protein kinases that exhibit nearly identical substrate specificities in vitro. Current paradigms hold that the relative contributions of ATM and ATR to nuclear substrate phosphorylation are dictated by the type of initiating DNA lesion; ATM-dependent substrate phosphorylation is principally activated by DNA double strand breaks, whereas ATR-dependent substrate phosphorylation is induced by UV light and other forms of DNA replication stress. In this report, we employed the cyclic AMP-response element-binding (CREB) protein to provide evidence for substrate discrimination by ATM and ATR in cellulo. ATM and ATR phosphorylate CREB in vitro, and CREB is phosphorylated on Ser-121 in intact cells in response to ionizing radiation (IR), UV light, and hydroxyurea. The UV light- and hydroxyurea-induced phosphorylation of CREB was delayed in comparison to the canonical ATR substrate CHK1, suggesting potentially different mechanisms of phosphorylation. UV light-induced CREB phosphorylation temporally correlated with ATM autophosphorylation on Ser-1981, and an ATM-specific small interfering RNA suppressed CREB phosphorylation in response to this stimulus. UV light-induced CREB phosphorylation was absent in ATM-deficient cells, confirming that ATM is required for CREB phosphorylation in UV irradiation-damaged cells. Interestingly, RNA interference-mediated suppression of ATR partially inhibited CREB phosphorylation in response to UV light, which correlated with reduced phosphorylation of ATM on Ser-1981. These findings suggest that ATM is the major genotoxin-induced CREB kinase in mammalian cells and that ATR lies upstream of ATM in a UV light-induced signaling pathway.     

Lysophosphatidic acid-induced c-fos up-regulation involves cyclic AMP response element-binding protein activated by mitogen- and stress-activated protein kinase-1

Lysophosphatidic acid (LPA) is a lipid growth factor that exerts diverse biological effects through its cognate receptor-mediated signaling cascades. Recently, we reported that LPA stimulates cAMP response element-binding protein (CREB) through mitogen- and stress-activated protein kinase-1 (MSK1). Previously, LPA has been shown to stimulate c-fos mRNA expression in Rat-2 fibroblast cells via a serum response element binding protein (SRF). However, involvement of CREB in LPA-stimulated c-fos gene expression is not elucidated yet. To investigate the CREB-mediated c-fos activation by LPA, various c-fos promoter-reporter constructs containing wild-type and mutated SRE and CRE were tested for their inducibility by LPA in transient transfection assays. LPA-stimulated c-fos promoter activation was markedly decreased when SRE and CRE were mutated. A dominant negative CREB significantly down-regulated the LPA-stimulated c-fos promoter activation. Chromatin immunoprecipitation assay revealed that LPA induced an increased binding of phosphorylated CREB and CREB-binding protein (CBP) to the CRE region of the endogenous c-fos promoter. Immunoblot analyses with various pharmacological inhibitors further showed that LPA induces up-regulation of c-fos mRNA level by activation of ERK, p38 MAPK, and MSK1. Taken together, our results suggest that CREB plays an important role in up-regulation of c-fos mRNA level in LPA-stimulated Rat-2 fibroblast cells.     

Regulation of cyclic AMP-dependent response element-binding protein (CREB) by the nociceptin/orphanin FQ in human dopaminergic SH-SY5Y cells

Nociceptin/orphanin FQ (N/OFQ), an endogenous ligand for opioid receptor-like (ORL1) receptor, transduces signaling cascades implicated in MAPK, PKC, PLC, and calcium, etc. This study was designed to investigate the intracellular signaling mechanism of N/OFQ in human dopaminergic neuroblastoma SH-SY5Y cells. N/OFQ rapidly induced the phosphorylation of CREB, which was significantly suppressed by pretreatment of PKA inhibitor, but not by MAPK inhibitors. It also time-dependently increased the phosphorylation of MAPK, which was proven as ERKs, whereas it did not affect the PI3K activity. Interestingly, KT5720, a specific inhibitor of PKA, markedly suppressed the phosphorylation of MAPK by N/OFQ in SH-SY5Y cells. Furthermore, BAPTA-AM, an intracellular chelator of Ca(2+), completely abolished the phosphorylation of CREB as well as MAPK in N/OFQ-treated SH-SY5Y cells. Taken together, these results suggest that N/OFQ independently induces the activation of CREB prior to MAPK phosphorylation, which was also modulated by PKA. Furthermore, Ca(2+)-related signaling implicates in the phosphorylation processes of CREB and MAPK simultaneously.