Samten Buka, Howard Susan T, Weis Steven E, Wu Shiping, Shams Homayoun, Townsend James C, Safi Hassan, Barnes Peter F
Center for Pulmonary and Infectious Disease Control, Department of Microbiology and Immunology, University of Texas Health Center, Tyler, TX 75708, USA.
J Immunol. 2005 May 15;174(10):6357-63. doi: 10.4049/jimmunol.174.10.6357.
IFN-gamma is essential for resistance to many intracellular pathogens, including Mycobacterium tuberculosis. Transcription of the IFN-gamma gene in activated T cells is controlled by the proximal promoter element (-73 to -48 bp). CREB binds to the IFN-gamma proximal promoter, and binding is enhanced by phosphorylation of CREB. Studies in human T cell lines and in transgenic mice have yielded conflicting results about whether CREB is a positive or a negative regulator of IFN-gamma transcription. To determine the role of CREB in mediating IFN-gamma production in response to a microbial pathogen, we evaluated the peripheral blood T cell response to M. tuberculosis in healthy tuberculin reactors. EMSAs, chromatin immunoprecipitation, and Western blotting demonstrated that stimulation of PBMC with M. tuberculosis induced phosphorylation and enhanced binding of CREB to the IFN-gamma proximal promoter. Neutralization of CREB with intracellular Abs or down-regulation of CREB levels with small interfering RNA decreased M. tuberculosis-induced production of IFN-gamma and IFN-gamma mRNA expression. In addition, M. tuberculosis-stimulated T cells from tuberculosis patients, who have ineffective immunity, showed diminished IFN-gamma production, reduced amounts of CREB binding to the IFN-gamma proximal promoter, and absence of phosphorylated CREB. These findings demonstrate that CREB positively regulates IFN-gamma production by human T cells that respond to M. tuberculosis.
干扰素-γ对于抵抗包括结核分枝杆菌在内的许多细胞内病原体至关重要。活化T细胞中干扰素-γ基因的转录受近端启动子元件（-73至-48碱基对）控制。CREB结合到干扰素-γ近端启动子上，且CREB的磷酸化可增强这种结合。关于CREB是干扰素-γ转录的正调节因子还是负调节因子，在人T细胞系和转基因小鼠中的研究结果相互矛盾。为了确定CREB在介导对微生物病原体应答中干扰素-γ产生的作用，我们评估了健康结核菌素反应者外周血T细胞对结核分枝杆菌的应答。电泳迁移率变动分析、染色质免疫沉淀和蛋白质免疫印迹表明，用结核分枝杆菌刺激外周血单核细胞可诱导CREB磷酸化并增强其与干扰素-γ近端启动子的结合。用细胞内抗体中和CREB或用小干扰RNA下调CREB水平可降低结核分枝杆菌诱导的干扰素-γ产生及干扰素-γ mRNA表达。此外，来自免疫无效的结核病患者的经结核分枝杆菌刺激的T细胞显示干扰素-γ产生减少、与干扰素-γ近端启动子结合的CREB量减少且不存在磷酸化的CREB。这些发现表明，CREB对响应结核分枝杆菌的人T细胞产生干扰素-γ起正调节作用。
