Molecular Genetic Analysis of Yeast TAFs

描述（由申请人提供）：我们研究的目的是阐明RNA聚合酶II控制真核mRNA产生的分子机制。mRNA充当蛋白质合成的蓝图或模板。mRNA和蛋白质生物合成的缺陷导致严重的人类疾病，如癌症、心血管疾病、糖尿病、出生缺陷和免疫缺陷。因此，如果我们要真正了解正常和病理状态，并最终干预治疗疾病，那么控制mRNA合成的过程的全面知识是必不可少的。我们的工作利用遗传上易于处理的面包酵母作为模型，用于阐明进化上高度保守的转录因子TFIID在RNA聚合酶II的mRNA合成或转录的复杂过程中所起的详细作用。TFIID由15个不同的蛋白质亚基组成，全TFIID复合物控制酵母和人类中超过90%的所有mRNA编码基因的转录。特别强调将放在研究两个转录因子，阻遏物激活蛋白1（Rap1p）和SBF，如何与TFIID合作，以调节编码核糖体蛋白编码基因（Rap1p）和细胞周期S期依赖的mRNA基因转录（SBF）的基因的激活。我们将使用多方面的方法，结合生物化学，生物物理，细胞生物学和遗传学的方法来检查这些转录因子相互之间的相互作用，基因DNA，基因特异性DNA结合调节因子Rap1p和SBF，以及实际合成mRNA的RNA聚合酶II酶，最后，作为适当的染色质修饰辅助调节因子。这些实验的成功完成将增加我们对mRNA基因转录机制的理解。鉴于所有这些调控过程在进化上都是保守的，我们在酵母模型系统中的工作将为人类mRNA基因转录控制机制提供见解，从而最终为人类疾病提供见解。

DESCRIPTION (provided by applicant): The goal of our research is to elucidate the molecular mechanisms controlling eukaryotic mRNA production by the enzyme RNA Polymerase II. mRNA serves as the blueprint, or template, for protein synthesis. Defects in mRNA and protein biosynthesis lead to serious human disease such as cancer, cardiovascular disease, diabetes, birth defects and immunodeficiency. Thus, a comprehensive knowledge of the processes controlling mRNA synthesis is essential if we are to truly understand both normal and pathological states, and ultimately intervene to cure disease. Our work utilizes the genetically tractable baker's yeast as a model for elucidating the detailed roles that the highly evolutionarily conserved transcription factor TFIID plays in the complex process of mRNA synthesis, or transcription, by RNA Polymerase II. TFIID is made up of 15 distinct protein subunits and the holo-TFIID complex controls the transcription of over 90% of all mRNA encoding genes in both yeasts and humans. Particular emphasis will be placed on studying how two transcription factors, Repressor Activator Protein 1 (Rap1p) and SBF, collaborate with TFIID to modulate activation of the genes encoding ribosomal protein-encoding genes (Rap1p) and cell-cycle S-phase-dependent mRNA gene transcription (SBF). We will use a multifaceted approach that combines biochemical, biophysical, cell biological and genetic methods to examine the interplay of these transcription factors with each other, gene DNA, the gene-specific DNA-binding regulatory factors Rap1p and SBF, and the RNA polymerase II enzyme that actually synthesizes mRNA, and finally, as appropriate chromatin-modifying coregulators. Successful completion of these experiments will increase our understanding mRNA gene transcription mechanisms. Given that all of these regulatory processes are evolutionarily conserved, our work in the yeast model system will provide insights into human mRNA gene transcription control mechanisms, and hence ultimately human disease.