Control of Transcriptional Attenuation of Stress-induced Genes in Yeast

描述(由申请人提供)：细胞存活取决于对细胞外环境压力信号的反应能力。不同的应激信号诱导特定基因的表达，这些基因在应激的生理反应中发挥作用。在没有压力的情况下，这些基因中的许多基因的表达都保持在最低水平。我们在酿酒酵母这个真核系统的模型中发现，许多应激诱导基因的基础表达通过一种新的机制-提前转录终止或转录减弱而被最小化。细胞壁应激诱导的基因需要MAP激酶Mpk1在转录过程中执行两个独立的步骤，这两个步骤都不需要它的蛋白激酶活性。第一种是招募一种转录因子作为目标基因的启动子。第二个涉及阻断衰减，这发生在非诱导条件下目标基因的启动子-近端区域。衰减是由Sen1终止复合体介导的，并被Mpk1到延伸RNA聚合酶(POL II)的易位所阻断。在诱导条件下，基因的表达依赖于衰减的缓解。对于Mpk1诱导的基因，这是通过Mpk1与延伸因子Paf1的关联发生的，Paf1阻止Sen1复合体招募到Pol II。这种相互作用在人类Mpk1的同源基因ERK5中保守，表明调节的转录衰减在人类中起作用。根据我们的初步发现，我们认为在非诱导条件下，许多胁迫诱导的基因都被转录衰减所沉默，并且一系列转录因子可能通过与Paf1C(一种含有Paf1的复合体)的相互作用来缓解诱导条件下的转录衰减。该项目的长期目标是提供一种新的方法，通过抑制转录衰减的缓解来阻断特定基因或基因组的表达。我们建议阐明调节转录衰减的机制，以及不同的胁迫在多大程度上利用相似或不同的衰减缓解因子来调节各种靶基因。一个直接的目标将是确定其他对不同信号做出反应的MAP激酶是否也起到了缓解衰减的作用。另一个项目将确定非MAPK衰减缓解因子，这些因子允许诱导我们发现处于衰减控制下的各种应激诱导基因。第三个目标是了解Paf1C在Sen1终止复合体被招募到Pol II中的作用。总体而言，这些研究将从机制上理解受调控的转录衰减，并揭示这一过程在酵母中的普遍存在，这将为后续的人类细胞研究提供信息。

DESCRIPTION (provided by applicant): Cell survival depends on the ability to respond to stress signals from the extracellular environment. Diverse stress signals induce the expression of specific genes that function in the physiologic response to the stress. In the absence of stress, expression of many of these genes is maintained at a minimal level. We have found in the yeast S. cerevisiae, a model eukaryotic system, that the basal expression of many stress-induced genes is minimized by a novel mechanism - premature transcriptional termination, or transcriptional attenuation. Genes induced by cell wall stress require the MAP kinase Mpk1 to carry out two separate steps in the transcription process, neither of which requires its protein kinase activity. The first is to recruit a transcription factor to promoters of target genes. The second involves blocking attenuation, which occurs within the promoter-proximal region of target genes under non-inducing conditions. Attenuation is mediated by the Sen1 termination complex and is blocked by the translocation of Mpk1 to the elongating RNA polymerase (Pol II). Under inducing conditions, gene expression depends upon the relief of attenuation. For Mpk1-induced genes, this happens through the association of Mpk1 with the elongation factor Paf1, which blocks the recruitment of the Sen1 complex to Pol II. This interaction is conserved in the human ortholog of Mpk1, ERK5, suggesting that regulated transcriptional attenuation operates in humans. Based on our preliminary findings, we propose that a wide variety of stress-induced genes are silenced by transcriptional attenuation under non-inducing conditions and that a constellation of transcription factors are likely to relieve attenuation under inducing conditions through interactions with the Paf1C (a complex containing Paf1). The long-term objective of this project is to provide a novel approach to blocking the expression of specific genes, or groups of genes, by inhibiting relief of transcriptional attenuation. We propose to elucidate the mechanisms that regulate transcriptional attenuation and the degree to which various stresses use similar or different attenuation-relief factors to regulate a variety of target genes. One immediate goal will be to determine if other MAP kinases that respond to different signals also function as attenuation-relief factors. Another project will identify non-MAP kinase attenuation- relief factors that allow the induction of a variety of stress-induced genes we have found to be under attenuation control. A third goal will be to understand the role of the Paf1C in the recruitment of the Sen1 termination complex to Pol II. Overall, these studies will yield a mechanistic understanding of regulated transcriptional attenuation and reveal the ubiquity of the process in yeast, which will inform subsequent studies on human cells.