Fitness Profile of HIV-1 Genome with Host Cofactor Selection Pressure

描述(由申请人提供)：拟议的研究将利用一种新的遗传平台，在选定的条件下对整个艾滋病毒基因组进行功能性分析，并在每个单个核苷酸位置发生突变。HIV基因组的遗传交互作用将由选定的宿主限制因子决定。该遗传平台将定量地识别 基因组(以及相应的编码氨基酸)是功能相互作用所必需的，代表着在识别宿主-病原体相互作用方面向前迈出的重要一步。使用研究小组开发并成功演示的新的高分辨率遗传图谱方法，这项研究将定量评估艾滋病毒-1基因组中每个核苷酸位置所有可能的核酸碱基变化的突变影响。饱和突变与下一代序列的结合将被应用于同时监测基因组所有位置的突变。在实现研究目标的过程中，这项研究的目标是建立衡量每个核苷酸位置对HIV-1复制的重要性的初始图谱，并随后确定基因组中对宿主限制因子干扰素诱导的跨膜蛋白3(IFITM3)的抗性和补偿效应的核苷酸位置。我们将使用一种特性良好的限制因子载脂蛋白B mRNA编辑酶--催化多肽样3G(APOBEC3G)作为对照。更重要的是，拟议的功能图谱方法将展示在单个实验平台上评估跨越整个病毒基因组的大型和多样化病毒突变群体参与复制过程或致病过程的能力，这不仅将影响艾滋病毒-1研究，而且将影响许多其他病毒病原体的研究。从获得的艾滋病毒-1全基因组的高分辨率图谱中，详细列出了所有可能的碱基变化在每个核苷酸位置的突变耐受性，这将证明对艾滋病毒/艾滋病疫苗的开发非常有益。病毒-细胞相互作用的识别提供了 为治疗性药物设计提供了大量潜在有用的新靶点。

DESCRIPTION (provided by applicant): The proposed research will utilize a novel genetic platform to functionally profile the entire HIV genome at selected conditions with mutation at every single nucleotide position. The genetic interaction of HIV genome will be determined with selected host restriction factors. The genetic platform will quantitatively identify regions of the genome (and the corresponding coded amino acids) that are essential for the functional interaction, and represents a major step forward to identify host-pathogen interactions. Using the novel high-resolution genetic profiling approach that the research group has developed and successfully demonstrated, the study will quantitatively assess mutational impact for all possible nucleic acid base changes for every nucleotide position within the HIV-1 genome. The combination of saturation mutagenesis with next generation sequence will be applied to monitor mutations at all positions of the genome simultaneously. In accomplishing the research goals, the study will aim to establish the initial profile gauging the essentialness of each nucleotide position for HIV-1 replication, and subsequently identify nucleotide positions in the genome for resistance and compensatory effects to host restriction factor interferon-induced trans-membrane protein 3 (IFITM3). A well-characterized restriction factor apolipoprotein B mRNA-editing enzyme, catalytic polypeptide- like 3G (APOBEC3G) will be used as a comparison. More importantly, the proposed functional profiling method will demonstrate the ability to evaluate a large and diverse viral mutant population spanning an entire viral genome in all genomic space for the involvement in a replication process or pathogenic process on a single experimental platform, which will impact not just HIV-1 research, but the study of many other viral pathogens. From the obtained high resolution map of the entire HIV-1 genome detailing mutational tolerability at each nucleotide position for all possible base changes, which should prove highly beneficial for HIV/AIDS vaccine development. Identification of viral-cellular interactions provides a great wealth of new potentially useful targets for therapeutic drug design.