CHEMOKINE RECEPTOR EXPRESSION IN LYMPHOCYTES

描述（改编自申请人摘要）：研究者 表明干扰素（IFN）上调了 趋化因子受体（CCR），CCR 1，CCR 3和CCR 5在原发性单核细胞源性 巨噬细胞（MDM）和单核细胞样细胞系U937中。 这种效果是 至少部分在RNA表达水平上介导， 涉及JAK-STAT信号通路。 自艾滋病毒进入目标以来 至少部分由趋化因子受体介导，因为免疫细胞 艾滋病的异常似乎包括抗原呈递缺陷， 由于艾滋病患者血清干扰素水平似乎升高， 干扰素介导的这些上调的机制分析 趋化因子受体似乎是必要的。 调查人员计划确定 通过克隆CCR 1、CCR 3和CCR 5启动子区的5'端， mRNA起始位点及其在瞬时转染中的表征 测定它们驱动报告基因表达的能力， 通过增加活性来响应IFNa和g。 由于5'端的 mRNA尚未明确确定，可能需要 通过5' RACE识别5'末端。 活性启动子的序列分析可 鉴定共有转录因子应答元件。 启动子 通过检测5 ′和3 ′缺失的活性可以更好地确定区域 变种人 转录因子的结合位点将通过 电泳迁移率变化和超迁移分析。 定点 诱变将被用来更好地确定结合位点 转录因子 这项工作可能会提供重要的线索， 趋化因子受体的表达受干扰素的影响， 影响感染者HIV-1复制的因素

DESCRIPTION (adapted from applicant's abstract): The investigators have shown that interferons (IFNs) upregulate the functional expression of chemokine receptors (CCR), CCR1, CCR3 and CCR5 in primary monocyte-derived macrophages (MDMs) and in the monocytoid cell line U937. This effect is mediated at least in part at the level of RNA expression, and it is likely to involve the JAK-STAT signaling pathway. Since entry of HIV into target cells is mediated at least in part by chemokine receptors, since the immune abnormalities in AIDS appear to include defects in antigen presentation, and since AIDS patients appear to have elevated serum levels of interferons, an analysis into the mechanisms of interferon-mediated upregulation of these chemokine receptors seems warranted. The investigators plan to identify the promoter regions of CCR1, CCR3 and CCR5 by cloning the regions 5' to the mRNA initiation sites and characterizing them in transient transfection assays for their ability to drive the expression of reporter genes and to respond to IFNa and g by an increase in activity. Since the 5' ends of the mRNAs have not been unambiguously determined, it may be necessary to identify the 5' ends by 5' RACE. Sequence analysis of active promoters may identify consensus transcription factor response elements. The promoter regions may be better defined by testing the activity of 5' and 3'deletion mutants. The binding sites for transcription factors will be identified by electrophoretic mobility shift and supershift assays. Site directed mutagenesis will be used to better define the binding sites of transcriptional factors. This work may provide important clues on how chemokine receptor expression is affected by IFNs and give insights on factors affecting HIV-1 replication in infected people.