A 68-nucleotide sequence within the 3′ noncoding region of simian hemorrhagic fever virus negative-strand RNA binds to four MA104 cell proteins
A 68-nucleotide sequence within the 3′ noncoding region of simian hemorrhagic fever virus negative-strand RNA binds to four MA104 cell proteins
复制标题
DOI：
10.1128/jvi.72.5.4341-4351.1998
复制
发表时间：
1998-05-01
影响因子：
5.4
通讯作者：
Brinton, MA
中科院分区：
文献类型：
--
作者：
Hwang, YK;Brinton, MA
猴出血热病毒（SHFV）的负链RNA [3 '（-）NCR RNA]的3'非编码区（NCR）长209个核苷酸（nt）。由于该3'区，命名为3'（-）209，是全长正链RNA的起始位点，并且是合成5'前导序列的模板，其存在于全长和亚基因组mRNA上，因此它可能含有RNA合成的顺式作用信号，并与细胞和病毒蛋白相互作用以形成复制复合物。凝胶迁移率变动分析表明，MA 104 S100细胞质提取物中的细胞蛋白与SHFV 3 '（-）209 RNA形成两种复合物，竞争凝胶迁移率变动分析的结果表明，这些相互作用是特异性的。四个蛋白质的分子量为103，86，55，和36 kDa的紫外线诱导的交联试验中检测到，这些蛋白质（103，55，和36 kDa）中的三个也被检测到西北印迹法。用未感染和感染的提取物获得了相同的凝胶迁移率变化和UV诱导的交联模式，表明检测到的四种蛋白是细胞蛋白，而不是病毒蛋白。四种细胞蛋白的结合位点定位于从SHFV负链RNA的3'端的nt 117和184之间的区域（68-nt序列）。预测该68-nt序列形成tao茎环、SL 4和SL 5。另一种动脉炎病毒，乳酸脱氢酶升高病毒C（LDV-C）的3 '（-）NCR RNA在竞争凝胶迁移率变化试验中与SHFV 3'（-）209 RNA竞争。W诱导的交联试验表明，与SHFV 3 '（-）209 RNA结合的分子量相同的四种MA 104细胞蛋白也与LDV-C 3'（-）NCR RNA和马动脉炎病毒3 '（-）NCR RNA结合。然而，这些病毒RNA中的每一种也结合到另外的MA 104蛋白。MA 104细胞蛋白的结合位点显示位于LDV-C 3 '（-）NCR和SHFV 3'（-）209 RNA中的相似位置。这些数据表明，一组细胞蛋白的结合位点在所有动脉炎病毒RNA中是保守的，并且这些细胞蛋白可用作病毒复制复合物的组分。
The 3' noncoding region (NCR) of the negative-strand RNA [3'(-)NCR RNA] of the arterivirus simian hemorrhagic fever virus (SHFV) is 209 nucleotides (nt) in length. Since this 3' region, designated 3'(-)209, is the site of initiation of full-length positive-strand RNA and is the template for the synthesis of the 5' leader sequence, which is found on both full-length and subgenomic mRNAs, it is likely to contain cis-acting signals for RNA synthesis and to interact with cellular and viral proteins to form replication complexes. Gel mobility shift assays showed that cellular proteins in MA104 S100 cytoplasmic extracts formed two complexes with the SHFV 3'(-)209 RNA, and results from competition gel mobility shift assays demonstrated that these interactions were specific. Four proteins with molecular masses of 103, 86, 55, and 36 kDa were detected in UV-induced cross-linking assays, and three of these proteins (103, 55, and 36 kDa) were also detected by Northwestern blotting assays. Identical gel mobility shift and UV-induced cross-linking patterns were obtained with uninfected and SHFV-infected extracts, indicating that the four proteins detected are cellular, not viral, proteins, The binding sites for the four cellular proteins were mapped to the region between nt 117 and 184 (68-nt sequence) from the 3' end of the SHFV negative-strand RNA. This 68-nt sequence was predicted to form tao stem-loops, SL4 and SL5. The 3'(-)NCR RNA of another arterivirus, lactate dehydrogenase-elevating virus C (LDV-C), competed with the SHFV 3'(-)209 RNA in competition gel mobility shift assays. W-induced cross-linking assays showed that four MA104 cellular proteins with the same molecular masses as those that bind to the SHFV 3'(-)209 RNA also bind to the LDV-C 3'(-)NCR RNA and equine arteritis virus 3'(-)NCR RNA. However, each of these viral RNAs also bound to an additional MA104 protein. The binding sites for the MA104 cellular proteins were shown to be located in similar positions in the LDV-C 3'(-)NCR and SHFV 3'(-)209 RNAs. These data suggest that the binding sites for a set of the cellular proteins are conserved in all arterivirus RNAs and that these cell proteins may be utilized as components of viral replication complexes.