PMID- 32917739 OWN - NLM STAT- MEDLINE DCOM- 20210712 LR - 20230829 IS - 1477-9137 (Electronic) IS - 0021-9533 (Linking) VI - 134 IP - 5 DP - 2020 Nov 9 TI - Cell-to-cell and genome-to-genome variability of adenovirus transcription tuned by the cell cycle. LID - jcs252544 [pii] LID - 10.1242/jcs.252544 [doi] AB - In clonal cultures, not all cells are equally susceptible to virus infection, and the mechanisms underlying this are poorly understood. Here, we developed image-based single-cell measurements to scrutinize the heterogeneity of adenovirus (AdV) infection. AdV delivers, transcribes and replicates a linear double-stranded DNA genome in the nucleus. We measured the abundance of viral transcripts using single-molecule RNA fluorescence in situ hybridization (FISH) and the incoming 5-ethynyl-2'-deoxycytidine (EdC)-tagged viral genomes using a copper(I)-catalyzed azide-alkyne cycloaddition (click) reaction. Surprisingly, expression of the immediate early gene E1A only moderately correlated with the number of viral genomes in the cell nucleus. Intranuclear genome-to-genome heterogeneity was found at the level of viral transcription and, in accordance, individual genomes exhibited heterogeneous replication activity. By analyzing the cell cycle state, we found that G1 cells exhibited the highest E1A gene expression and displayed increased correlation between E1A gene expression and viral genome copy numbers. The combined image-based single-molecule procedures described here are ideally suited to explore the cell-to-cell variability in viral gene expression in a range of different settings, including the innate immune response. CI - (c) 2020. Published by The Company of Biologists Ltd. FAU - Suomalainen, Maarit AU - Suomalainen M AUID- ORCID: 0000-0002-3276-3008 AD - Department of Molecular Life Sciences, University of Zurich, 8057 Zurich, Switzerland. FAU - Prasad, Vibhu AU - Prasad V AUID- ORCID: 0000-0001-7450-607X AD - Department of Molecular Life Sciences, University of Zurich, 8057 Zurich, Switzerland. FAU - Kannan, Abhilash AU - Kannan A AUID- ORCID: 0000-0002-8502-4129 AD - Department of Molecular Life Sciences, University of Zurich, 8057 Zurich, Switzerland. FAU - Greber, Urs F AU - Greber UF AUID- ORCID: 0000-0003-2278-120X AD - Department of Molecular Life Sciences, University of Zurich, 8057 Zurich, Switzerland urs.greber@mls.uzh.ch. LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20201109 PL - England TA - J Cell Sci JT - Journal of cell science JID - 0052457 SB - IM MH - *Adenoviridae/genetics MH - Cell Cycle/genetics MH - Genome, Viral/genetics MH - In Situ Hybridization, Fluorescence MH - *Virus Replication/genetics OTO - NOTNLM OT - Cell cycle OT - Cell-to-cell variability OT - DNA virus OT - Nucleus OT - Phenotypic heterogeneity OT - Single mRNA OT - Single virus genome OT - Transcription COIS- Competing interestsThe authors declare no competing or financial interests. EDAT- 2020/09/13 06:00 MHDA- 2021/07/13 06:00 CRDT- 2020/09/12 05:29 PHST- 2020/08/03 00:00 [received] PHST- 2020/08/27 00:00 [accepted] PHST- 2020/09/13 06:00 [pubmed] PHST- 2021/07/13 06:00 [medline] PHST- 2020/09/12 05:29 [entrez] AID - jcs.252544 [pii] AID - 10.1242/jcs.252544 [doi] PST - epublish SO - J Cell Sci. 2020 Nov 9;134(5):jcs252544. doi: 10.1242/jcs.252544.