ISOLATION AND ANALYSIS OF MURINE LEUKEMIA VIRUS RECEPTOR

小鼠C型逆转录病毒嗜亲性宿主范围的测定 通过表达特定的病毒受体。 这个目标 我们的建议是了解病毒受体的分子基础， 在感染中发挥作用。 具体目标是： 1. 完成一个完整的嗜亲性病毒受体的克隆 基因 完整基因在非允许性人EJ中的表达 细胞将赋予亲嗜性宿主范围。 2. 确定受体的表达是否是唯一的决定因素 通过将受体基因引入广泛的嗜亲性宿主范围， 真核生物物种的范围和感染测试。 3. 研究受体结构之间的关系 和功能 a. 确定转染的鼠基因是否编码 直接或通过指定受体的修饰， 内源性人类蛋白质。 B. 将编码区的核苷酸序列与 序列库，以确定受体基因是否已被 被鉴定为细胞蛋白质。 C. 模型受体结构相对于膜。 确定可能参与包膜的胞外结构域 约束力 D. 通过制备抗合成的抗血清来测试该模型。 来自预测的胞外结构域的肽。 1. 测试这些抗血清阻断包膜结合的能力， 感染 2. 证明病毒包膜蛋白之间的直接相互作用 （gp70）和受体的免疫共沉淀。 3. 研究膜gp70-受体相互作用在 干扰 e. 确定病毒与受体相互作用的特定位点。 方法是在绑定中创建无效突变， 然后选择突变病毒， 补体 这些研究将有助于阐明 逆转录病毒宿主范围，逆转录病毒进入的机制， 细胞和干扰现象。此外，他们将 鉴定被逆转录病毒利用的细胞蛋白， 生命周期中的重要一步。 在人类逆转录病毒中， 相互作用已被证明在 艾滋病和逆转录病毒引起的癌症的发病机制。

Ecotropic host range in murine type C retroviruses is determined by the expression of a specific virus receptor. The goal of this proposal is to understand the molecular basis for viral receptor function in infection. The specific aims are to: 1. Complete the cloning of an intact ecotropit virus receptor gene. Expression of an intact gene in non-permissive human EJ cells will confer ecotropic host range. 2. Determine if receptor expression is the sole determinant of ecotropic host range by introducing the receptor gene into a broad range of eucaryotic species and testing for infection. 3. Investigate the relationship between the receptor structure and function. a. Determine if the transfected murine gene encodes the receptor directly or by specifying the modification of an endogenous human protein. b. Compare the nucleotide sequence of the coding region to a sequence bank to determine if the receptor gene has been identified as a cellular protein. c. Model receptor structure with respect to the membrane. Identify extracellular domain(s) that may be involved in envelope binding. d. Test this model by preparing antisera against synthetic peptides from the predicted extracellular domains. 1. Test the ability of these antisera to block envelope binding and infection. 2. Demonstrate direct interaction between virus envelope protein (gp70) and receptor by co-immunoprecipitation. 3. Investigate the role of membrane gp70-receptor interaction in interference. e. Identify specific sites of virus envelope-receptor interaction. The approach will be to create null mutations in the binding domain of the receptor and then to select mutant viruses that complement. These studies will help elucidate the molecular determinants of retrovirus host range, the mechanism of retrovirus entry into cells, and the phenomenon of interference. Further, they will identify a cellular protein that is utilized by retroviruses for an essential step in their lifecycle. In human retroviruses these interactions have been shown to play an important role in the pathogenesis of AIDS and retrovirus-induced cancer.