Applequist S E, Selg M, Raman C, Jäck H M
Department of Microbiology and Immunology, Stritch School of Medicine, Loyola University of Chicago, Maywood, IL 60153, USA.
Nucleic Acids Res. 1997 Feb 15;25(4):814-21. doi: 10.1093/nar/25.4.814.
Levels of most nonsense mRNAs are normally reduced in prokaryotes and eukaryotes when compared with that of corresponding functional mRNAs. Genes encoding polypeptides that selectively reduce levels of nonsense mRNA have so far only been identified in simple eukaryotes. We have now cloned a human cDNA whose deduced amino acid sequence shows the highest degree of homology to that of UPF1, a bona fide Saccharomyces cerevisiae group I RNA helicase required for accelerated degradation of nonsense mRNA. Based on the total sequence of the shorter yeast UPF1 protein, the overall identity between the human protein and UPF1 is 51%. Besides NTPase and other RNA helicase consensus motifs, UPF1 and its human homolog also share similar putative zinc finger motifs that are absent in other group I RNA helicases. Northern blot analysis with the human cDNA probe revealed two transcripts in several human cell lines. Further, antibodies raised against a synthetic peptide of the human polypeptide detected a single 130 kDa polypeptide on Western blots from human and mouse cells. Finally, immunofluorescence and Western blot analyses revealed that the human and mouse polypeptides, like yeast UPF1, are expressed in the cytoplasm, but not in the nucleus. We have thus identified the first mammalian homolog of yeast UPF1, a protein that regulates levels of nonsense mRNA, and we tentatively name this protein human HUPF1 (for human homolog of UPF1).
与相应的功能性mRNA相比，大多数无义mRNA的水平在原核生物和真核生物中通常会降低。到目前为止，编码能选择性降低无义mRNA水平的多肽的基因仅在简单真核生物中被鉴定出来。我们现已克隆出一种人类cDNA，其推导的氨基酸序列与UPF1的氨基酸序列具有最高程度的同源性，UPF1是酿酒酵母中一种真正的I类RNA解旋酶，是加速无义mRNA降解所必需的。根据较短的酵母UPF1蛋白的完整序列，人类蛋白与UPF1之间的总体一致性为51%。除了NTPase和其他RNA解旋酶共有基序外，UPF1及其人类同源物还共享相似的假定锌指基序，而这些基序在其他I类RNA解旋酶中不存在。用人cDNA探针进行的Northern印迹分析在几种人类细胞系中检测到两种转录本。此外，针对人类多肽的合成肽产生的抗体在来自人类和小鼠细胞的Western印迹上检测到一条单一的130 kDa多肽。最后，免疫荧光和Western印迹分析表明，人类和小鼠多肽与酵母UPF1一样，在细胞质中表达，但不在细胞核中表达。我们因此鉴定出了酵母UPF1的首个哺乳动物同源物，一种调节无义mRNA水平的蛋白，我们暂定将该蛋白命名为人类HUPF1（UPF1的人类同源物）。
