HIV-1 host cell factors: a saturation screen for human chromosomes 4, 6, 21 and X

描述（由申请人提供）：HIV-1宿主细胞因子：人类染色体4、6、21和x的饱和筛选。人们对鉴定调节HIV-1生命周期的细胞宿主细胞因子有浓厚的兴趣。这些因素可分为抑制感染的限制因素和帮助感染过程的允许因素。这些因素的研究将有助于我们理解病毒和宿主细胞的相互作用，并确定新的细胞药物靶点。由于目前的药物治疗是基于靶向病毒蛋白，而HIV的高突变率能够对这些抗病毒药物产生耐药性，因此细胞蛋白可能构成更难产生耐药性的靶点。已经使用了各种策略来识别这些蛋白质。在这里，我们提出了一种鉴定HIV早期生命周期宿主细胞因子的方法。我们的目标是做一个饱和插入突变筛选，将取样1/5的人类基因组的宿主细胞因子基因。我们将使用具有4、6、21和x染色体单体的细胞来实现这一目标。我们将开发一种破坏基因功能的插入载体，并使用一种对感染有毒性的HIV-1载体选择感染了这种诱变载体的细胞来抵抗HIV-1感染。插入染色体4、6、21和X将通过突变载体两侧的DNA测序来鉴定。在这项研究结束时，我们将至少有2-3个经过验证的宿主细胞因子和它们在生命周期中被利用的点。我们也将拥有一个宝贵的资源——宿主细胞因子的敲除细胞系。这将有助于在未来的研究中了解蛋白质和病毒的结构-功能关系。这些宿主细胞因子可能构成小分子药物对抗HIV-1和艾滋病的新靶点。

DESCRIPTION (provided by applicant): HIV-1 host cell factors: a saturation screen for human chromosomes 4, 6, 21 and X. There is intense interest in identifying cellular host cell factors that modulate the life cycle of HIV-1. These factors can be classified as restriction factors that inhibit infection and permissive factors that aid the infection process. The study of these factors will aid in our understanding of viral and host cell interactions and to identify novel cellular drug targets. Since present drug therapies are based on targeting viral proteins, and the high mutation rate of HIV is able to generate resistance to these antiviral drugs, cellular proteins may constitute targets in which resistance is harder to generate. Various strategies to identify these proteins have been used. Here we propose an approach for the identification of host cell factors for the early life cycle of HIV. We aim to do a saturating insertional mutagenesis screen that will sample 1/5 of the human genome for host cell factor genes. We will achieve this aim using cells that have monosomies of chromosomes 4, 6, 21 and X. We will develop an insertional vector that disrupts gene function and select cells infected with this mutagenic vector for resistance to HIV-1 infection using an HIV-1 vector that is toxic on infection. Insertions into chromosome 4, 6, 21 and X will be identified through sequencing of DNA flanking the mutagenic vector. At the conclusion of this study we will have at least 2-3 verified host cell factors and the point in the lifecycle at which they are utilized. We will also have in hand a valuable resource - a knockout cell line of the host cell factor. This will enable structure-function relationships of the protein and the virus in future studies. These host cell factors may constitute novel targets for small drug molecules to combat HIV-1 and AIDS. PUBLIC HEALTH RELEVANCE: HIV-1 host cell factors: a saturation screen for human chromosomes 4, 6, 21 and X. HIV-1 relies on host cell proteins to complete its replication cycle. Identification of these factors is important since they are potential targets for the development of drugs to combat HIV-1 infection. In this study we aim to interrogate 1/5 the human genome in a pilot study to discover some host cell factors required by HIV-1 for infection.