S CEREVISIAE GENERAL TRANSCRIPTION FACTOR TFIID

这个子项目是许多利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可在其他CRISP条目中表示。所列机构为 中心，但不一定是研究者所在的机构。 转录因子TFIID是一种多亚基复合物，负责将RNA聚合酶II和其他相关因子的组装靶向并成核至真核生物中的启动子起始位点。它对基因表达的调节很重要。基因表达的错误调节可导致发育缺陷、代谢紊乱和癌症。了解转录起始机制是一个重要目标，有关TFIID的结构信息将有助于实现这一目标。估计TFIID复合物含有1.1-1.2 MDa的蛋白质，由15个单独的亚基贡献。由于分离的亚基的表达和溶解性问题以及难以获得足够的材料，这种大复合物的结构信息一直难以获得。我们的目标是扩展由P. Schultz小组使用串联亲和纯化（TAP-标签）分离材料进行的酵母TFIID复合物的负染色电子显微镜研究。我们将获得TFIID复合体的冷冻电镜重建。这些数据将用于我们获得高分辨率X射线晶体结构的努力。使用冷冻EM进一步表征TFIID与其他已知基础因子的相互作用，以进一步揭示启动子处RNA聚合酶II转录系统各个组分之间的空间关系，这将是特别有趣的。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Transcription factor TFIID is a multi-subunit complex responsible for targeting and nucleating the assembly of RNA polymerase II and other associated factors to promoter start sites in eukaryotic organisms. It is important for the regulation of gene expression. Mis-regulation of gene expression can cause developmental defects, metabolic disorders, and cancer. Understanding the mechanism of transcriptional initiation is an important goal, and structural information about TFIID would contribute towards this end. The TFIID complex is estimated to contain 1.1-1.2 MDa of protein contributed from 15 individual subunits. Structural information for this large complex has been difficult to obtain due to expression and solubility problems for the isolated subunits and difficulty in obtaining sufficient material. Our goal has been to extend on negative stain electron microscopy studies of the yeast TFIID complex carried out by the group of P. Schultz using Tandem Affinity Purification (TAP-tag) isolated material. We will obtain cryo-EM reconstructions of the TFIID complex. This data will be used in our efforts to obtaining a high-resolution X-ray crystal structure. It would be of particular interest to further characterize the interactions of TFIID with other known basal factors using cryo-EM to further reveal the spatial relationships between the various components of the RNA Polymerase II transcription system at promoters.