FUNCTION OF CLATHRIN-COATED MEMBRANES

笼状蛋白包被的囊泡和膜与多种 真核细胞的细胞内转运过程。精准的 将讨论笼蛋白在细胞生长和蛋白质转运中的作用。 氯化菊酯重油生产缺陷酵母突变株的鉴定 和轻亚基。用多克隆抗血清鉴定了一个克隆。 酵母重链基因(CHC1)在Lambdagtll基因组中的表达 图书馆。通过杂交选择确认了该插入片段的同源性 翻译重链mRNA后进行免疫沉淀。一个 然后使用插入片段来生成删除的 染色体CHC1基因座。令人惊讶的是，可行的缺失突变菌株 生产的产品没有可检测到的免疫反应重链，并且 产生的囊泡也缺乏网状蛋白轻链。虽然是笼状结构 缺失突变细胞生长缓慢，分泌糖蛋白转移酶 几乎是正常的。现在将检查缺失突变细胞的比率 以及蛋白质到不同目的地的运输和分类的保真度。 将利用敏感的核寻找网状蛋白的结构类似物 酸杂交程序和通过检查替代关联 在重链缺失突变细胞中形成网状蛋白轻链。 轻链基因的克隆和缺失分析将用于 检测轻链在蛋白质运输中可能发挥的任何独立作用。

Clathrin-coated vesicles and membranes have been implicated in a variety of intracellular transport processes in eukaryotic cells. The precise funciton of clathrin in cell growth and protein transport will be addressed by evaluation of yeast mutants defective in production of clathrin heavy and light subunits. Polyclonal antiserum was used to identify a clone of the yeast heavy chain gene (CHC1) expressed in a Lambdagtll genomic library. The identity of this insert was confirmed by hybrid-selected translation of heavy chain mRNA followed by immune precipitation. A fragment of the insert was then used to generate deletions of the chromosomal CHC1 locus. Surprisingly, viable deletion mutant strains were produced which have no detectable immunoreactive heavy chain, and which produce vesicles that also lack clathrin light chain. Although clathrin deletion mutant cells grow slowly, secretion of the glycoprotein invertase is nearly normal. Deletion mutant cells will now be examined for the rate and fidelity of transport and sorting of proteins to various destinations. Structural analogues of clathrin will be sought using sensitive nucleic acid hybridization procedures and by examining alternative associations formed with clathrin light chain in heavy chain deletion mutant cells. Cloning and deletion analysis of the light chain gene will be used to detect any independent role that light chains may play in protein transport.