Qu X, Yang Z, Zhang S, Shen L, Dangel A W, Hughes J H, Redman K L, Wu L C, Yu C Y
Children's Hospital Research Foundation, Molecular, Cellular and Developmental Biology Program, Department of Pediatrics, The Ohio StateUniversity, Columbus, OH 43205-2696, USA.
Nucleic Acids Res. 1998 Sep 1;26(17):4068-77. doi: 10.1093/nar/26.17.4068.
The human helicase gene SKI2W is located between RD and RP1 in the class III region of the major histocompatibility complex. Transcripts of SKI2W are detectable in RNA samples isolated from multiple tissues. The protein product Ski2w shares striking amino acid sequence similarities to the yeast antiviral protein Ski2p that controls the translation of mRNAs, probably based on the mRNA structural integrity. Whether this translational regulation mechanism for cellular and viral RNAs exists in mammals is under investigation. Antisera against human Ski2w were generated using fusion proteins produced in bacteria or insect cells. Western blot analysis showed that the endogenous Ski2w protein is approximately 140 kDa in size and is enriched in polysomal fractions of cytoplasmic extracts from HeLa cells. Ribosomal profile studies revealed that Ski2w distributed throughout the entire sucrose gradient in the presence of Mg2+, but co-sedimented with the 18S rRNA-containing 40S subunit and the small ribosomal subunit protein S27a in the presence of EDTA. The co-sedimentation of Ski2w with the 40S subunit is not affected by RNase A treatment of the cell extract, or the addition of KCl to 0.5 M, suggesting that Ski2w is associated with the 40S ribosomal subunit. Indirect immunofluorescence experiments showed that human Ski2w is localized in the nucleoli and in the cytoplasm. In essence, human Ski2w is present at the sites of ribosome biogenesis and protein synthesis.
人类解旋酶基因SKI2W位于主要组织相容性复合体III类区域的RD和RP1之间。在从多个组织分离的RNA样本中可检测到SKI2W的转录本。蛋白质产物Ski2w与控制mRNA翻译的酵母抗病毒蛋白Ski2p具有显著的氨基酸序列相似性,这可能基于mRNA的结构完整性。这种针对细胞和病毒RNA的翻译调控机制在哺乳动物中是否存在正在研究中。使用在细菌或昆虫细胞中产生的融合蛋白制备了针对人类Ski2w的抗血清。蛋白质印迹分析表明,内源性Ski2w蛋白大小约为140 kDa,在HeLa细胞胞质提取物的多核糖体组分中富集。核糖体图谱研究表明,在Mg2+存在下,Ski2w分布在整个蔗糖梯度中,但在EDTA存在下,与含18S rRNA的40S亚基和小核糖体亚基蛋白S27a共沉降。Ski2w与40S亚基的共沉降不受细胞提取物的RNase A处理或添加KCl至0.5 M的影响,表明Ski2w与40S核糖体亚基相关。间接免疫荧光实验表明,人类Ski2w定位于核仁和细胞质中。本质上,人类Ski2w存在于核糖体生物发生和蛋白质合成的部位。