ANALYSIS OF INTRACELLULAR SMALL RNAS OF MOUSE HEPATITIS-VIRUS - EVIDENCE FOR DISCONTINUOUS TRANSCRIPTION
ANALYSIS OF INTRACELLULAR SMALL RNAS OF MOUSE HEPATITIS-VIRUS - EVIDENCE FOR DISCONTINUOUS TRANSCRIPTION
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DOI：
10.1016/0042-6822(87)90414-4
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发表时间：
1987-02-01
期刊：
影响因子：
3.7
通讯作者：
LAI, MMC
中科院分区：
文献类型：
--
作者：
BARIC, RS;SHIEH, CK;LAI, MMC
我们之前已经证明在小鼠肝炎病毒（MHV）感染的细胞中存在多种小的含先导RNA物种。在本文中，我们分析了这些小rna的起源、结构和合成机制。利用特异性先导RNA和基因A、D和F的cDNA探针，我们证明了这些小RNA的亚群来源于各种病毒基因。这些亚群具有离散和可重复的大小，随其衍生的基因而变化。每个子集的大小与二级结构区域相关，其自由能范围为-1.6至-77.1 kcal/mol，在所检查的每个mrna中。此外，在复制中间体（RI） RNA上检测到相同的亚群，表明它们代表功能性转录中间体。这些小rna的生物学意义得到了进一步的支持，检测到长度为47、50和57个核苷酸的含先导rna，它们对应于两种MHV重组病毒的交叉位点。这些数据，再加上MHV感染期间RNA重组的高频率，表明病毒聚合酶可能在二级结构区域或周围暂停，从而产生大量含有游离先导的RNA中间体，这些中间体可以与模板重新结合，作为合成全长RNA或重组RNA的引物。这些数据表明MHV转录使用一种不连续和非过程机制，其中RNA聚合酶允许部分RNA产物在转录过程中暂时与模板分离。
We have previously shown the presence of multiple small leader-containing RNA species in mouse hepatitis virus (MHV)-infected cells. In this paper, we have analyzed the origin, structure, and mechanism of synthesis of these small RNAs. Using cDNA probes specific for leader RNA and genes A, D, and F, we demonstrate that subsets of these small RNAs were derived from the various viral genes. These subsets have discrete and reproducible sizes, varying with the gene from which they are derived. The size of each subset correlates with regions of secondary structure, whose free energy ranges from -1.6 to -77.1 kcal/mol, in each of the mRNAs examined. In addition, identical subsets were detected on the replicative intermediate (RI) RNA, suggesting that they represent functional transcriptional intermediates. The biological significance of these small RNAs is further supported by the detection of leader-containing RNAs of 47, 50, and 57 nucleotides in length, which correspond to the crossover sites in two MHV recombinant viruses. These data, coupled with the high frequency of RNA recombination during MHV infection, suggest that the viral polymerase may pause in or around regions of secondary structure, thereby generating pools of free leader-containing RNA intermediates which can reassociate with the template, acting as primers for the synthesis of full-length or recombinant RNAs. These data suggest that MHV transcription uses a discontinuous and nonprocessive mechanism in which RNA polymerase allows the partial RNA products to be dissociated from the template temporarily during the process of transcription.