Chen Z, Faaberg K S, Plagemann P G
Department of Microbiology, University of Minnesota, Minneapolis 55455.
Virus Res. 1994 Nov;34(2):167-77. doi: 10.1016/0168-1702(94)90098-1.
The mechanism of synthesis of the seven subgenomic mRNAs of lactate dehydrogenase-elevating virus (LDV) was explored. One proposed mechanism, leader-primed transcription, predicts the formation of free 5'-leader in infected cells which then primes reinitiation of transcription at specific complementary sites on the antigenomic template. No free LDV 5'-leader of 156 nucleotides was detected in LDV-infected macrophages. Another mechanism, independent replication of the subgenomic mRNAs, predicts the presence of negative complements to all subgenomic mRNAs in infected cells which might be generated from subgenomic mRNAs in virions. Full-length antigenomic RNA was detected in LDV-infected macrophages by Northern hybridization at a level of < 1% of that of genomic RNA, but no negative polarity subgenomic RNAs. Negative complements to all subgenomic mRNAs, however, were detected by reverse transcription of total RNA from infected macrophages using as primer an oligonucleotide complementary to the antileader followed by polymerase chain reaction amplification using this sense primer in combination with various oligonucleotide primers complementary to a segment downstream of the junction between the 5' leader and the body of each subgenomic RNA. It is unclear whether these minute amounts of negative subgenomic RNAs function in the replication of the subgenomic mRNAs. They could also be by-products of the RNA replication process. Finally, no subgenomic mRNAs were detected in LDV virions.
对乳酸脱氢酶升高病毒（LDV）七种亚基因组mRNA的合成机制进行了探索。一种提出的机制，即前导引物转录，预测在受感染细胞中会形成游离的5'-前导序列，然后该序列在反基因组模板上的特定互补位点引发转录重新起始。在LDV感染的巨噬细胞中未检测到156个核苷酸的游离LDV 5'-前导序列。另一种机制，即亚基因组mRNA的独立复制，预测在受感染细胞中存在所有亚基因组mRNA的负链互补序列，这些序列可能由病毒粒子中的亚基因组mRNA产生。通过Northern杂交在LDV感染的巨噬细胞中检测到全长反基因组RNA，其水平低于基因组RNA的1%，但未检测到负极性亚基因组RNA。然而，使用与反前导序列互补的寡核苷酸作为引物，对来自感染巨噬细胞的总RNA进行逆转录，然后使用该正义引物与各种与每个亚基因组RNA的5'前导序列和主体之间连接处下游片段互补的寡核苷酸引物进行聚合酶链反应扩增，从而检测到所有亚基因组mRNA的负链互补序列。尚不清楚这些微量的负链亚基因组RNA是否在亚基因组mRNA的复制中起作用。它们也可能是RNA复制过程的副产物。最后，在LDV病毒粒子中未检测到亚基因组mRNA。
