THE ROLE OF INTRACELLULAR TRAFFIC IN HIV INFECTION

核膜的运输在几个步骤中是必要的。 艾滋病毒的生命周期。一旦进入细胞质，病毒RNA就会转化为 必须进入细胞核的双链DNA。病毒监管 蛋白质进入细胞核，而病毒转录本被输出到 细胞质。病毒Rev蛋白已被确定为一种关键的调节因子 HIV包膜基因的运输。已经采取了几种方法来 了解Rev的工作原理。正在准备的细胞系 表达进入细胞核的REV/糖皮质激素受体嵌合体 一种可诱导的方式；这些细胞系还包含一种结构，其中包含 REV响应元件，用于控制 氯霉素乙酰基转移酶(CAT)基因。这将允许CAT 可以方便地跟随允许直接测量出口的表达式 CAT m RNA进入细胞质。还设计了一种方法来 在外源添加的DNA周围体外生成细胞核。该方法使用 非洲爪哇卵的提取物。聚集在这种提取物中的核 以多种方式模拟相间核，实现活化核 运输。这些准备工作使检查REV的机制成为可能 参与艾滋病毒生命周期的其他分子的行动和运动。 在其他研究中，已经检查了核孔的结构。这个 核孔需要糖蛋白成分才能正常形态和 功能。以往的研究表明核孔的过度表达 糖蛋白可能对细胞有毒性。为了绕过这些问题， 编码主要核糖蛋白p62的cdna已放置在 糖皮质激素反应性表达载体的控制。通过表达 正义和反义结构，应该有可能研究 这种核孔成分的抑制或过度表达的影响。 孔道复合体的许多成分现在已经是分子 克隆的。了解逆转录病毒产品如何穿过核膜 对于试图调节或抑制 艾滋病毒的生命周期。

Transport across the nuclear membrane is necessary at several steps in the life cycle of HIV. Once in the cytoplasm, the viral RNA is converted to double stranded DNA which must enter the nucleus. Viral regulatory proteins enter the nucleus while viral transcripts are exported into the cytoplasm. The viral REV protein has been identified as a key regulator of transport of HIV envelope mRNA. Several approaches have been taken to understand how REV may function. Cell lines are being prepared which express a REV/glucocorticoid receptor chimera which enters the nucleus in an inducible fashion; these cell lines also contain a construct containing a REV response element controlling expression of a chloramphenicolacetyltransferase (CAT) gene. This will allow CAT expression to be conveniently followed allowing a direct measure of export of CAT mRNA into the cytoplasm. A means has also been devised for generating nuclei in vitro around exogenously added DNA. The method uses extracts from Xenopus laevis eggs. The nuclei assembled in such extracts mimic interphase nuclei in many ways and carry out active nuclear transport. These preparations allow examination of the mechanism of REV action and the movement of other molecules involved in the HIV life cycle. In other studies, the structure of the nuclear pore has been examined. The nuclear pore requires glycoprotein components for proper morphology and function. Previous studies indicate the overexpression of nuclear pore glycoproteins may be toxic to cells. To circumvent these problems, the cDNA encoding the major nuclear glycoprotein p62 has been placed under the control of a glucocorticoid responsive expression vector. By expressing the sense and antisense constructs, it should be possible to study the effects of suppression or overexpression of this nuclear pore component. A number of the components of the pore complex have now been molecularly cloned. Understanding how retroviral products cross the nuclear envelope is critical to attempts to regulate or inhibit the critical steps in the HIV life cycle.