Temporal Patterns of Human Cytomegalovirus Transcription: Mapping the Viral RNAs Synthesized at Immediate Early, Early, and Late Times After Infection

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Abstract
The transcription of the human cytomegalovirus genome was investigated at immediate early, early and late times after infection. Viral RNAs associated with either the whole cell, the nucleus, the cytoplasm or the polyribosomes were analyzed. At immediate early times, i.e., in the absence of de novo viral protein synthesis, the viral RNA in high abundance originated from a region of the long unique section of the prototype arrangement of the viral genome (0.660-0.770 map units). The viral RNA in low abundance originated from the long repeat sequences (0.010-0.035 and 0.795-0.825 map units) and a region in the long unique section (0.201-0.260 map units). Viral RNA associated with the polyribosomes as polyadenylated RNA were mapped to these restricted regions of the viral genome and characterized according to size class in kilobases. At 24 h after infection in the presence of an inhibitor of viral DNA replication, i.e., at early times, the stable viral RNA in highest abundance mapped in the long repeat sequences. Viral RNAs at intermediate abundance mapped in 2 regions of the long unique section of the viral genome (0.325-0.460 and 0.685-0.770 map units). Stable viral RNA that were associated with the polyribosomes in high abundance as polyadenylated RNA originated from the long repeat sequences but not from the long unique section of the viral genome. An analysis of whole-cell RNA at late times (72 h) indicated that the abundant transcription was in the regions of the long unique sequences (0.325-0.460 and 0.660-0.685 map units), and transcription of intermediate abundance was from the long repeat sequences were associated with the polyribosomes at late times after infection. mRNA originating from the long and short unique sequences were found associated with the polyribosomes at higher relative concentration than at early times after infection. Expression of the immediate early viral genes apparently is required to transcribe the early viral genes in the long repeat and adjacent sequences. These sequences are also transcribed at late times after infection while viral DNA synthesis continues. The expression of viral genes in most of the long and short unique sequences appears to require viral DNA replication. [Human fibroblast cells were used in this study.].