pUL79 alters the rate of transcriptional elongation at viral loci.

<p>(A) Schematic representation of the HCMV genes and host genes examined by the nuclear run-on (NRO) assay. Locations and sizes of primer-probe pairs used in subsequent RT-qPCR analysis are indicated. (B–D) HFFs were infected with AD<i>dd</i>UL79 at an MOI of 3 in the presence or absence of 1 µM Shld1 (indicated by “+” or “−” sign, respectively). Nuclear extracts were prepared at 24 or 72 hpi and analyzed by NRO assays. Transcription elongation was allowed to resume for 30 minutes in the presence of biotin-labeled UTP, labeled RNA was isolated, and their amounts were determined by RT-qPCR. In addition, accumulations of total RNAs were also determined by RT-qPCR. The normalized amounts of viral run-on transcripts or total transcripts in the presence of Shld1 were set at 1 for the NRO assay or total transcript accumulation analysis, respectively. (B) Relative amounts of total and run-on transcripts of viral late genes UL99 and UL32. (C) Relative amounts of run-on and total transcripts of viral immediate-early and early genes. (D) Relative amounts of run-on and total transcripts of host genes. Data from three independent experiments were collected and standard deviations were calculated by Prism 6 software. Statistical analysis was performed using Student's <i>t</i> test (**, P<0.01; ***, P<0.005; NS, not significant).