Long G4 regions (LG4s) in the human genome constitute functional enhancers that coordinate neighboring gene expressions

人类基因组中的长 G4 区域 (LG4) 构成协调邻近基因表达的功能增强子

• 批准号: 2223547
• 负责人: Glen Borchert
• 金额: $ 42.73万
• 依托单位: University of South Alabama
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-15 至 2024-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2223547&HistoricalAwards=false
• 关键词: Long; G4; regions; LG4s; human

A promoter is a region of DNA upstream of a gene where proteins bind to initiate transcription of (turn on) a gene by producing an RNA copy of the gene sequence. Promoters can be very complex and typically work together with other DNA regions known as enhancers to ensure specific, robust gene transcription. Fortunately, despite their complexity, promoters are easy to identify because they generally correspond to the 100-1,000 base pairs (bp) just before the start of a gene. This has allowed detailed characterization of a massive number of promoters over the past several decades. However, enhancer characterizations have proven much more challenging. Like promoters, enhancers are typically short, 100-1,000 bp DNA sequences that markedly regulate gene transcription but, unlike promoters, an enhancer can be found up to 1 million bp away from the gene it influences. How do enhancers work across such large distances? Genomic DNA can bend, bringing enhancers and promoters close to one another to initiate transcription. Interestingly, numerous studies attempting to link genomic mutations with various traits and/or diseases have found that more than 90% of disease-associated mutations lie within the noncoding portions of the genome, and even more strikingly, most of these mutations lie within regions of the genome thought to function as enhancers rather than promoters. Despite their importance, there is still only a very limited ability to define enhancer sequences, and major questions about their mechanisms of action and targeting determinants remain unanswered. Understanding what drives enhancer:promoter interaction can significantly improve our ability to address a number of societal challenges (e.g. disease, improving crop resistance to adverse environmental conditions via gene modulation, etc.). This project also provides unique interdisciplinary training for students (including local underrepresented minority students) combining molecular biology and computational genetics/bioinformatics.A better understanding of the interplay between gene promoters and gene-distal enhancers will unquestionably lead to fundamental new insights into the basic control of gene transcription and the action of regulatory mutations involved in disease. However, the fundamental mechanisms responsible for how enhancers target and transfer information to their cognate promoters, broadly speaking, remain key gaps in our knowledge. The work outlined in this proposal will directly address the long-standing question of how enhancers, which are often located far (hundreds of kilobase pairs) away, make contact with specific promoters in the context of the nucleus. Notably, enhancers are often significantly enriched with G4-capable sequences and, interestingly, the promoters regulated by G4-enhancers are likewise disproportionately enriched with G4-capable sequences. As hybrid G4 structures can be formed between distinct DNA strands, together, these observations suggest that direct G4:G4 interactions can contribute to enhancer:promoter targeting. The work outlined in this proposal is significant in that it will employ an array of genetic manipulations, biochemical assays, and novel NGS-IP strategies to directly characterize the role of G4 DNA in enhancer::promoter targeting and generate novel methodologies that will be of use in defining specific enhancer::promoter interactions.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.

启动子是基因上游的DNA区域，蛋白质在这里结合，通过产生基因序列的RNA拷贝来启动基因的转录（开启）。启动子可能非常复杂，通常与其他DNA区域（称为增强子）一起工作，以确保特定的、健壮的基因转录。幸运的是，尽管它们很复杂，但启动子很容易识别，因为它们通常对应于基因开始前的100- 1000个碱基对（bp）。在过去的几十年里，这使得对大量启动子的详细描述成为可能。然而，增强子的表征已被证明更具挑战性。与启动子一样，增强子通常是短的100-1,000 bp的DNA序列，可以显著调节基因转录，但与启动子不同的是，增强子可以在距离其影响基因100万bp的地方找到。增强剂是如何在如此大的距离上起作用的？基因组DNA可以弯曲，使增强子和启动子彼此靠近，从而启动转录。有趣的是，许多试图将基因组突变与各种特征和/或疾病联系起来的研究发现，超过90%的与疾病相关的突变位于基因组的非编码部分，更引人注目的是，这些突变中的大多数位于被认为是增强子而不是启动子的基因组区域。尽管它们很重要，但定义增强子序列的能力仍然非常有限，关于它们的作用机制和靶向决定因素的主要问题仍然没有答案。了解是什么驱动增强子：启动子相互作用可以显著提高我们应对许多社会挑战的能力（例如疾病，通过基因调节提高作物对不利环境条件的抗性等）。该项目还为学生（包括本地少数族裔学生）提供独特的跨学科培训，结合分子生物学和计算遗传学/生物信息学。更好地了解基因启动子和基因远端增强子之间的相互作用，无疑将导致对基因转录的基本控制和涉及疾病的调节突变作用的根本新见解。然而，从广义上讲，增强子如何靶向并将信息传递给其同源启动子的基本机制仍然是我们知识中的关键空白。本提案中概述的工作将直接解决一个长期存在的问题，即通常位于很远（数百千碱基对）的增强子如何与细胞核中的特定启动子接触。值得注意的是，增强子通常富含G4-capable序列，有趣的是，由g4增强子调控的启动子同样不成比例地富含G4-capable序列。由于混合G4结构可以在不同的DNA链之间形成，这些观察结果表明，G4:G4的直接相互作用可以促进增强子：启动子的靶向。本提案中概述的工作具有重要意义，因为它将采用一系列遗传操作，生化分析和新型NGS-IP策略来直接表征G4 DNA在增强子：：启动子靶向中的作用，并产生将用于定义特定增强子：：启动子相互作用的新方法。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。