From genomics to proteomics

所有生物体都不断暴露于造成DNA损伤的环境因素，包括来自阳光的紫外线、辐射和绝大多数（约90%）化学致癌物，这对物种的生存和公众健康构成极大威胁。我们的研究重点是这些药物如何导致基因组不稳定、细胞死亡和癌症，以及生物体如何应对这些攻击并保护自己。我实验室的主要研究重点是研究最近描述的真核生物从简单酵母到人类的DNA损伤耐受性（DDT）途径。我们对参与这一途径的关键基因的初步发现做出了贡献，这一途径不会消除DNA损伤，而是允许复制机制绕过障碍，无论是否引起突变。由于其历史短，运作复杂，直到最近人们才知道这种途径是如何运作的。本质上，真核生物中的DDT是通过称为PCNA的复制钳的共价修饰来调节的。特别地，单个泛素（Ub）的附着可导致致突变旁路，而与Ub链的进一步附着可导致非致突变旁路；后者如何实现尚不清楚。我的实验室最近发现，一种叫做9-1-1的类似pcna的夹子，专门用于感知和响应DNA损伤，也可以被Ub修饰，这种修饰在细胞对DNA损伤的耐受性中起着关键作用。我们的中心目标是研究细胞如何通过在酵母模型、培养的人类细胞以及动物模型中泛素化两个DNA夹来协调滴滴涕过程。最近，我们开始将注意力从遗传转向生化方法，旨在了解控制各种滴滴涕信号通路的详细分子事件。在我们提出的每一个研究项目中，都有很大一部分致力于蛋白质的生化表征。因此，所要求的色谱系统将在确保我们的研究计划的成功发挥至关重要的作用。

All living organisms are constantly exposed to environmental agents that cause DNA damage, including UV from sunlight, radiation and the vast majority (>90%) of chemical carcinogens, which pose a great danger to the survival of species and public health. Our research focuses on how these agents cause genomic instability, cell death and cancer, and how living organisms respond to these assaults and protect themselves. The major focus of research in my laboratory is to study a recently described pathway called DNA damage tolerance (DDT) in eukaryotes, from simple yeast to human. We contributed to the initial discovery of key genes involved in this pathway, which does not remove DNA damage, but instead allows the replication machinery to bypass the roadblock with or without causing mutations. Due to its short history and complexity in operation, little was known about how this pathway operated until recently. In essence, DDT in eukaryotes is regulated through covalent modifications of a replication clamp called PCNA. In particular, attachment by a single ubiquitin (Ub) leads to a mutagenic bypass, whereas further attachment with a Ub chain leads to non-mutagenic bypass; how the latter is achieved remains unclear. My laboratory recently discovered that a PCNA-like clamp called 9-1-1, which is dedicated to sensing and responding to DNA damage, can also be modified by Ub and that this modification plays a key role in cellular tolerance to DNA damage. Our central objective is to investigate how cells coordinate the DDT process by ubiquitinating two DNA clamps in the model yeast, in cultured human cells, as well as in an animal model. Recently, we began to shift our attention from genetic to biochemical approaches aiming at understanding detailed molecular events governing various DDT signaling pathways.  In each of our proposed research projects, a significant portion is dedicated to biochemical characterization of proteins.  Hence, the requested chromatographic system will play a vital role in ensuring the success of our research programs.