TRANSPORT OF VIRAL PROTEINS IN MAMMALIAN CELLS

哺乳动物细胞中病毒蛋白的运输

• 批准号: 3293617
• 负责人: JOSEPH F SAMBROOK
• 金额: $ 20.95万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-05-01 至 1991-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3293617
• 关键词: cell nucleus; exocytosis; gene expression; glycosylation; hamsters; membrane permeability; membrane proteins; microorganism hemagglutinin; mutant; protein sequence; protein signal sequence; simian virus 40; tissue mosaicism; transport proteins; virus antigen; virus protein

The general aim of our laboratory is to understand the mechanisms which govern the direction and speed with which viral proteins expressed in mammalian cells are transported from their sites of synthesis to their final destinations. We are approaching this problem in two ways (i) by analyzing a series of genetically-engineered mutant and chimeric viral proteins that are defective at different stages of their particular transport pathways (ii) by isolating mutants of mammalian cells that are defective in transporting viral proteins to specific destinations. Recently, two types of amino acid sequences have been shown to be necessary and sufficient to guide proteins to these very different cellular locations. One of these - hydrophobic signal sequences - causes nascent polypeptides to be translocated across the membrane of the rough endoplasmic reticulum, from where they may travel to the Golgi apparatus and then to the cell surface or to cellular vesicular cytoplasmic organelles such as lysosomes. The other, more recently discovered, causes proteins to be transported to and retained in the cell nucleus. These karyophilic signals, like hydrophobic signals, vary greatly in sequence from protein to protein; in their simplest forms they consist of a short tract of largely basic amino acids. The goal of this application is to understand how two viral proteins (SV40 T antigen and influenze virus hemagglutinin) move to their specific destinations: the oncogenic SV40 T antigen is transported both to the nucleus and the plasma membrane of transformed cells. Influenze virus hemagglutinin (HA) is a prototypical integral membrane protein that moves with high efficiency along the conventional exocytotic pathway to the cell surface. In particular we will (i) isolate and characterize cellular mutants that are unable to transport HA to the cell surface along the conventional exocytotic pathway (ii) investigate the properties and intracellular transport of a set of chimeric SV40 T antigens and contain a hydrophobic sequence derived from another protein (influenza virus hemagglutinin) at their N-terminus (iii) test the hypothesis that SV40 T antigen associated with the plasma membrane reaches the cell surface, not by the conventional secretory pathway, but by transport through the cell nucleus (iv) analyze the transport of HA along the exocytotic pathway in lines of mutant CHO cells that display grossly altered patterns of glycosylation.

我们实验室的总体目标是了解 控制着病毒蛋白表达的方向和速度， 哺乳动物细胞从它们的合成位点运输到它们的 最终目的地。 我们从两个方面处理这个问题：（i） 分析一系列基因工程突变体和嵌合病毒 蛋白质在其特定的不同阶段有缺陷， （ii）通过分离哺乳动物细胞的突变体， 在将病毒蛋白质运送到特定目的地方面有缺陷。 最近，两种类型的氨基酸序列已被证明是必要的 足以引导蛋白质进入这些不同的细胞 地点 其中之一-疏水信号序列-导致新生的 多肽被转运穿过粗糙的膜 内质网，从那里他们可以前往高尔基体 然后转移到细胞表面或细胞泡状细胞质 细胞器如溶酶体。 另一个是最近发现的， 蛋白质被运送到细胞核并保留在细胞核中。 这些 亲核信号，像疏水信号一样，在序列上变化很大 从蛋白质到蛋白质;在它们最简单的形式中，它们由一个短的 主要是碱性氨基酸的一段。 本申请的目的是了解两种病毒蛋白（SV 40） T抗原和流感病毒血凝素）移动到其特异性 目的地：致癌SV 40 T抗原被转运到 转化细胞的细胞核和质膜。 流感病毒 血凝素（HA）是一种典型的整合膜蛋白， 沿着常规的细胞外吞途径高效地进入细胞 面 特别是，我们将（i）分离和表征细胞 不能将HA沿细胞表面沿着转运至细胞表面的突变体， 传统胞吐途径（ii）研究其性质并 一组嵌合SV 40 T抗原的细胞内转运，并含有 衍生自另一种蛋白质（流感病毒）疏水序列 （iii）检验SV 40 T 与质膜相关的抗原到达细胞表面，而不是 通过传统的分泌途径，而是通过细胞运输 （iv）分析HA沿着胞吐途径在细胞核中的转运 突变CHO细胞系显示出明显改变的 糖基化