Collaborative Research: NSF/MCB-BSF: The effect of transcription factor binding on UV lesion accumulation

人类基因组的DNA序列因人而异。然而，这些变异或突变并不均匀地分布在整个基因组中。当调控蛋白与DNA结合时，变异最大，尽管这些变化可以显著影响人类生物学，但人们对这一过程知之甚少。这个项目将确定基因组变异增加是因为蛋白质结合使DNA更容易损伤并随后发生突变，还是因为蛋白质阻止了受损DNA的修复导致突变。评估这些可能性需要开发创新的方法。这个项目不仅将回答蛋白质是否会使DNA更容易受损或更难修复的问题，而且新开发的方法将是DNA损伤定量测量方面的重大飞跃。为了确保新开发的方法产生更广泛的影响，研究人员将通过两种方式接触下一代科学家。将为高中生举办一个名为《是什么在破坏我的DNA》的研讨会，以吸引年轻一代的参与。此外，还将为需要接受处理更大数据集的培训的学术科学家提供一个名为“学术专业人员的计算思维”的密集夏季研讨会。转录因子(TF)蛋白质与整个基因组中的特定DNA序列结合，以调节基因表达。令人惊讶的是，最近的基因组研究表明，Tf结合位点的特征是DNA损伤增加，并且高度富含体细胞突变，这表明Tf-DNA结合具有诱变性。该项目假设，转录因子在结合部位诱导独特的损伤累积模式，这是由转录因子促进损伤形成和阻断核苷酸切除修复机制的能力所决定的。紫外光将被用作DNA损伤的来源，以发展高通量的体外技术和定量模型：来自不同家族的转录因子与含有损伤的DNA的亲和力和与修复蛋白的竞争能力；以及通过TF结合对DNA损伤形成的调控。这些模型将用于预测人类细胞的全基因组DNA损伤和修复，并开发基于细胞的基因组损伤和修复分析。这项工作的成功完成将把TF结合位点突变率的增加与TF增加人类基因组中紫外线损伤频率的分子机制联系起来。这项美国和以色列的合作项目得到了美国国家科学基金会的支持，联合资金来自分子和细胞生物科学、已建立的刺激竞争研究计划(EPSCoR)和以色列双国科学基金会。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

The DNA sequence of the human genome varies among people. However, these variations or mutations are not evenly dispersed throughout the genome. Variation is highest where regulatory proteins bind to DNA, a process that is poorly understood despite the fact that these changes can significantly impact human biology. This project will determine whether genomic variation is increased because protein binding makes DNA more susceptible to damage, and subsequently mutations, or because proteins block repair of damaged DNA leading to mutations. Assessing these possibilities requires development of innovative methods. This project will not only answer the question of whether proteins make DNA more susceptible to damage or harder to repair, but the newly developed methods will be a major leap forward in quantitative measurement of DNA damage. To ensure the broader impact of the newly developed methods, investigators will reach out to the next generations of scientists in two ways. A workshop entitled “What is Damaging My DNA” will be offered to high school students to engage the younger generation. In addition, an intensive summer workshop entitled “Computational Thinking for Academic Professionals” will be offered to academic scientists who need training handling larger datasets, such as those in this project.Transcription factor (TF) proteins bind to specific DNA sequences across the genome to regulate gene expression. Surprisingly, recent genomic studies indicate that TF binding sites are characterized by increased DNA damage and are highly enriched for somatic mutations, suggesting that TF‐DNA binding is mutagenic. This project hypothesizes that that TFs induce unique patterns of lesion accumulation within binding sites, which are dictated by the ability of TFs to enhance lesion formation and to occlude access to the nucleotide excision repair machinery. UV light will be used as a source of DNA damage to develop high‐throughput in vitro techniques and quantitative models for: the affinity of TFs from different families for lesion-containing DNA and ability to compete with repair proteins; and the modulation of DNA damage formation by TF binding. These models will be used to predict genome‐wide DNA damage and repair in human cells and to develop cell‐based assays of damage and repair in genomic contexts. Successful completion of this work will link increases in mutation rates in TF binding sites to the molecular mechanisms by which TFs increase UV damage frequency in the human genome.This collaborative US/Israel project is supported by the US National Science Foundation, with joint funding from Molecular and Cellular Biosciences and the Established Program to Stimulate Competitive Research (EPSCoR), and the Israeli Binational Science Foundation.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.