THE ROLE OF THE NUCLEAR ENVELOPE IN INTRACELLULAR PROTEIN SORTING

穿过核孔复合体的运输是一个必不可少的过程 调节细胞生长和正常发育。的结构。 正在研究核孔及其在核运输中的作用 一个分子水平。此前，该实验室已经表明，核子 毛孔复合体是由一系列具有共价的糖蛋白组成的。 连接O-连接的N-乙酰氨基葡萄糖。主核的糖基化 对孔糖蛋白p62进行了详细的研究。大鼠的基因和基因 编码RAT p62的基因已被分离出来。初级序列的检验 揭示了一系列由14个重复的五肽基序组成的具有 对GFSFG进行测序。抗该五肽序列的兔抗血清 不仅与p62反应，而且与核孔的其他成员反应。 糖蛋白家族。这些抗血清已被用来分离cDNA 编码其他核孔糖蛋白。此外，还会退化 对应于五肽重复的寡核苷酸已被用作 利用聚合酶链法分离额外的cdna物种的引物 反应技术。检查核孔的功能 核转运中的糖蛋白，体外核组装和 已经启用了运输系统。当将DNA添加到黄瓜提取物中时 非洲爪哇的核改造，核膜和核完整 毛孔。这些提取物可以耗尽内源性核孔。 组分，并用重组或生化改变的孔进行重组 蛋白质。利用这一分析方法显示了核孔的组装和运输 需要核孔糖蛋白；O-连接的N-乙酰氨基葡萄糖 部分可以在不改变运输的情况下进行修改。产生的能力 其他核孔糖蛋白的重组形式应该允许 进一步剖析这些核孔的功能作用 组件。

Transport across the nuclear pore complex is an essential process for regulating cell growth and normal development. The structure of the nuclear pore and its involvement in nuclear transport are being studied at a molecular level. Previously the laboratory has shown that the nuclear pore complex is made up of a family of glycoproteins having covalently attached O-linked N-acetylglucosamine. Glycosylation of the major nuclear pore glycoprotein p62 has been examined in detail. The rat cDNA and gene encoding rat p62 have been isolated. Examination of the primary sequence of p62 revealed a series of 14 repeating pentapeptide motifs having the sequence GFSFG. Rabbit antisera raised against this pentapeptide sequence react not only with p62 but also with the other members of the nuclear pore glycoprotein family. These antisera have been used to isolate the cDNAs encoding other nuclear pore glycoproteins. In addition, degenerate oligonucleotides corresponding to the pentapeptide repeat have been used as primers for isolating additional cDNA species using the polymerase chain reaction technique. To examine the function of the nuclear pore glycoproteins in nuclear transport, an in vitro nuclear assembly and transport system has been employed. When DNA is added to extracts of Xenopus laevis nuclei reform with intact nuclei envelopes and nuclear pores. These extracts can be depleted of endogenous nuclear pore components and reconstituted with recombinant or biochemically altered pore proteins. Using this assay nuclear pore assembly and transport were shown to require nuclear pore glycoproteins; the O-linked N-acetylglucosamine moiety can be modified without altering transport. The ability to generate recombinant forms of other nuclear pore glycoproteins should allow a further dissection of the functional roles of these nuclear pore components.