Frequency, magnitude, and mechanisms of collateral fitness effects of mutations

完全了解突变的影响对于理解生物学和疾病的治疗非常重要。有害突变通常被理解为通过破坏突变蛋白质最初应该做的事情来引起负面影响。 然而，突变的有害影响也可能来自于导致突变蛋白质做它不应该做的事情，例如干扰无关的生物过程。这些被称为附带效应。关于突变在多大程度上通过后一种机制造成有害影响，人们知之甚少。 在这项研究中，研究人员将全面量化两种抗生素耐药蛋白突变的附带影响，并揭示突变效应的机制。这将有助于理解蛋白质如何进化以及突变影响细胞生理学的机制，这对于理解正常的生物学功能以及人类疾病和抗生素耐药性的进化非常重要。每年的这项研究，两名研究生，两名本科生，一名高中生将在研究培训。 研究生将参加科学会议，介绍他们的研究，并获得指导本科生和高中生的指导经验。 突变的附带适应性效应是指那些不是来自基因/蛋白质执行其生理功能的能力变化的效应。突变对蛋白质进化的附带适应性效应的贡献是未知的。在这项研究中，将使用大规模诱变、生长竞争和深度测序技术的组合来测量两种抗生素耐药蛋白中所有单错义突变和所有单密码子插入/缺失突变的附带适应性效应。这些数据还将用于识别突变，以测试附带适应性效应背后机制的假设。 例如，由于蛋白质进化速率与蛋白质表达水平反相关，一种可能性是当突变导致蛋白质错误折叠或与其他蛋白质错误相互作用时，会发生有害的附带适应性效应。关于附带适应性效应的频率、大小和机制知之甚少，但这些信息对于理解突变适应性效应的分布和起源以及解释蛋白质进化的速率至关重要。对两种蛋白质中的这种效应进行系统和全面的研究将产生丰富的数据，从中可以了解附带适应性效应对突变适应性效应分布和蛋白质进化速率的贡献。该奖项反映了NSF的法定使命，并被认为值得通过使用基金会的智力价值和更广泛的影响审查标准进行评估来支持。

A complete understanding of the effects of mutations is important for understanding biology and the treatment of disease. Deleterious mutations are generally understood to cause negative effects by disrupting what the mutated protein was originally supposed to do. However, the deleterious effects of mutations might also arise from causing the mutated protein to do things it should not do, such as interfere with unrelated biological process. These are known as collateral effects. Very little is known about the extent to which mutations cause deleterious effects by the latter mechanism. In this research, the investigators will comprehensively quantify the collateral effects of mutations in two antibiotic resistance proteins and uncover mechanisms for the mutational effects. This will contribute to the understanding of how proteins evolve and the mechanisms by which mutations can affect the physiology of the cell, which is important for understanding normal biological function as well as human disease and the evolution of antibiotic resistance. Each year of this research, two graduate students, two undergraduates, and one high school student will be trained in research. The graduate students will attend scientific conferences to present their research and gain mentorship experience supervising the undergraduate and high school students. Collateral fitness effects of mutations are those effects which do not derive from changes in the ability of the gene/protein to perform its physiological functions. The contribution of collateral fitness effects of mutations to protein evolution is unknown. In this research, collateral fitness effects of all single missense mutations and all single-codon insertion/deletion mutations in two antibiotic resistance proteins will be measured using a combination of large scale mutagenesis, growth competition, and deep sequencing technology. The data will also be used to identify mutations to test hypotheses of the mechanisms behind collateral fitness effects. For example, since protein evolution rates anti-correlate with protein expression levels, one possibility is that deleterious collateral fitness effects occur when mutations cause the protein to misfold or misinteract with other proteins. Very little is known about the frequency, magnitude, and mechanisms of collateral fitness effects, but such information is critical for understanding the distribution and origins of fitness effects of mutations and interpreting the rates of protein evolution. The systematic and comprehensive study of such effects in two proteins will generate a wealth of data from which to understand the contribution of collateral fitness effects to the distribution of fitness effects of mutations and to protein evolution rates.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.