Tollervey D, Lehtonen H, Carmo-Fonseca M, Hurt E C
EMBL, Heidelberg, Germany.
EMBO J. 1991 Mar;10(3):573-83. doi: 10.1002/j.1460-2075.1991.tb07984.x.
The yeast snoRNP protein, NOP1, is structurally and functionally homologous to vertebrate fibrillarin and is essential for viability. A conditionally lethal allele was constructed by placing NOP1 expression under the control of a GAL promoter. Growth on glucose medium results in the depletion of NOP1 over several generations, during which cell growth is progressively impaired. Pulse labelling of proteins shows that NOP1 depleted strains are greatly impaired in the production of cytoplasmic ribosomes, and they have a reduced level of rRNA. Northern hybridization and pulse-chase labelling of pre-rRNA show a progressive impairment of all pre-rRNA processing steps. The pathway leading to 18S rRNA is particularly affected. Methylation of pre-rRNA is concomitantly impaired and unmethylated pre-rRNA accumulates and is not processed over long periods. NOP1 depletion does not prevent the accumulation of seven snoRNAs tested including U3; the levels of two species, U14 and snR190, decline. The snoRNAs synthesized in the absence of NOP1 retain TMG cap structures. Subnuclear fractionation and immunocytochemistry indicate that they continue to be localized in the nucleolus.
酵母小核仁核糖核蛋白（snoRNP）蛋白NOP1在结构和功能上与脊椎动物的纤维蛋白原同源，是细胞存活所必需的。通过将NOP1的表达置于GAL启动子的控制之下构建了一个条件致死等位基因。在葡萄糖培养基上生长会导致几代细胞中NOP1的耗尽，在此期间细胞生长逐渐受到损害。蛋白质脉冲标记显示，NOP1耗尽的菌株在细胞质核糖体的产生方面受到极大损害，并且它们的rRNA水平降低。前体rRNA的Northern杂交和脉冲追踪标记显示所有前体rRNA加工步骤都逐渐受到损害。导致18S rRNA的途径受到的影响尤为明显。前体rRNA的甲基化同时受到损害，未甲基化的前体rRNA积累，并且长时间不进行加工。NOP1的耗尽并不妨碍所测试的七种小核仁RNA（snoRNA）（包括U3）的积累；U14和snR190这两种小核仁RNA的水平下降。在没有NOP1的情况下合成的小核仁RNA保留了三甲基鸟苷（TMG）帽结构。亚核分级分离和免疫细胞化学表明它们继续定位于核仁中。
