Wente S R, Blobel G
Laboratory of Cell Biology, Howard Hughes Medical Institute, Rockefeller University, New York 10021.
J Cell Biol. 1994 Jun;125(5):955-69. doi: 10.1083/jcb.125.5.955.
We have isolated and characterized the gene encoding a fourth yeast glycine-leucine-phenylalanine-glycine (GLFG) repeat nucleoporin with a calculated molecular mass of 145.3 kD, and therefore termed NUP145. The amino-terminal half of Nup145p is similar to two previously identified GLFG nucleoporins, Nup116p and Nup100p (Wente, S. R., M. P. Rout, and G. Blobel. 1992. J. Cell Biol. 119:705-723). A deletion/disruption in the amino-terminal half of NUP145 (nup145 delta N) had only a slight effect on cell growth at temperatures between 17 and 37 degrees C. However, immunofluorescence microscopy of nup145 delta N cells with antinucleoporin antibodies showed that the characteristic punctate nuclear staining normally seen in wild-type yeast cells was reduced, with the majority of the signal located in one or two intense spots at the nuclear periphery. Thin section electron microscopy analysis revealed the presence of what appeared to be successive herniations of the nuclear envelope forming grape-like structures at primarily one site on the nup145 delta N nuclei. These successive herniations contained numerous NPC-like structures, correlating to the limited bright patches of anti-nucleoporin immunofluorescence signal. In some cases the successive herniations were small. Occasionally, however, multi-lobulated nuclei were seen. We suggest that the ultrastructural phenotype of nup145 delta N cells is due to a defective interaction of nup145 delta N NPCs with the surrounding pore membrane domain of the nuclear envelope. We have also analyzed the synthetic lethal phenotypes among GLFG nucleoporin mutant alleles, and found that strains harboring nup116 and either nup100 or nup145 mutations were not viable. This, in combination with the morphological analysis, may reflect overlapping yet distinct roles for these three GLFG nucleoporins in NPC-nuclear envelope interactions.
我们已经分离并鉴定了编码第四种酵母甘氨酸 - 亮氨酸 - 苯丙氨酸 - 甘氨酸（GLFG）重复核孔蛋白的基因，其计算分子量为145.3 kD，因此命名为NUP145。Nup145p的氨基末端一半与之前鉴定的两种GLFG核孔蛋白Nup116p和Nup100p相似（温特，S.R.，M.P.劳特，和G.布洛贝尔。1992。《细胞生物学杂志》119：705 - 723）。NUP145氨基末端一半的缺失/破坏（nup145 delta N）在17至37摄氏度之间的温度下对细胞生长只有轻微影响。然而，用抗核孔蛋白抗体对nup145 delta N细胞进行免疫荧光显微镜观察发现，野生型酵母细胞中通常可见的特征性点状核染色减少，大部分信号位于核周边的一两个强烈斑点处。超薄切片电子显微镜分析显示，在nup145 delta N细胞核的主要一个位点上，存在似乎是核膜连续疝出形成葡萄状结构的情况。这些连续的疝出包含许多类似核孔复合体的结构，与抗核孔蛋白免疫荧光信号的有限明亮斑块相关。在某些情况下，连续的疝出较小。然而，偶尔也会看到多叶核。我们认为nup145 delta N细胞的超微结构表型是由于nup145 delta N核孔复合体与核膜周围孔膜结构域的相互作用缺陷所致。我们还分析了GLFG核孔蛋白突变等位基因之间的合成致死表型，发现携带nup116以及nup100或nup145突变的菌株无法存活。这与形态学分析相结合，可能反映了这三种GLFG核孔蛋白在核孔复合体 - 核膜相互作用中重叠但又不同的作用。
