{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,1]],"date-time":"2026-07-01T02:39:50Z","timestamp":1782873590994,"version":"3.54.5"},"update-to":[{"DOI":"10.1371\/journal.ppat.1013374","type":"new_version","label":"New version","source":"publisher","updated":{"date-parts":[[2025,8,8]],"date-time":"2025-08-08T00:00:00Z","timestamp":1754611200000}}],"reference-count":51,"publisher":"Public Library of Science (PLoS)","issue":"8","license":[{"start":{"date-parts":[[2025,8,4]],"date-time":"2025-08-04T00:00:00Z","timestamp":1754265600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/publicdomain\/zero\/1.0\/"}],"funder":[{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["R21 AI130453"],"award-info":[{"award-number":["R21 AI130453"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["5T32AI007343-35"],"award-info":[{"award-number":["5T32AI007343-35"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000738","name":"U.S. Department of Veterans Affairs","doi-asserted-by":"publisher","award":["Merit award I01 BX004434 and SeqCure Network"],"award-info":[{"award-number":["Merit award I01 BX004434 and SeqCure Network"]}],"id":[{"id":"10.13039\/100000738","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["www.plospathogens.org"],"crossmark-restriction":false},"short-container-title":["PLoS Pathog"],"abstract":"<jats:p>Cell type differences in the human cytomegalovirus (HCMV) transcriptome may arise from variations in transcription or post-transcription regulation. Here we report unexpected differences in transcription and epigenetic control in late-stage HCMV infection of human differentiated NTera2 neural lineage cells (D-NT2) compared to fibroblasts, using integrated functional genomic approaches (PRO-Seq, RNA-Seq, DNA fragmentation factor-ChIP Seq, rapid viral protein degradation, and promoter mutation and function assays). In D-NT2, but not fibroblasts, RNA polymerase II initiation and elongation at several viral promoters requires viral DNA synthesis and are independent of host P-TEFb, viral immediate-early protein 2 (IE2), or viral late transcription factor (LTF). This includes transcription from the enhancer for the major immediate early (MIE) promoter where GC-box sequence mutations increase enhancer transcription, while mutations in CREB and NF-<jats:sc>kB<\/jats:sc> response elements reduce it. The GC-box mutations also alter infected D-NT2 cell morphology and gene expression program without affecting viral MIE gene expression levels, whereas mutations in CREB and NF-<jats:sc>k<\/jats:sc>B response elements do not induce these changes. In D-NT2, LTF-driven promoters constitute a smaller proportion of the viral late promoter population and are generally less active. Additionally, viral genomes have more nucleosomes, potentially restricting LTF access. A TATA-binding protein (TBP)-IE2-nucleosome complex, with more nucleosome than in fibroblasts, occupies the MIE promoter transcription start site, potentially contributing to its epigenetic silencing.<\/jats:p>","DOI":"10.1371\/journal.ppat.1013374","type":"journal-article","created":{"date-parts":[[2025,8,4]],"date-time":"2025-08-04T17:39:42Z","timestamp":1754329182000},"page":"e1013374","update-policy":"https:\/\/doi.org\/10.1371\/journal.ppat.corrections_policy","source":"Crossref","is-referenced-by-count":1,"title":["Cell type differences in human cytomegalovirus transcription and epigenetic regulation with insights into major immediate-early enhancer-promoter control"],"prefix":"10.1371","volume":"21","author":[{"given":"Qiaolin","family":"Hu","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ming","family":"Li","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mrutyunjaya","family":"Parida","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Benjamin M.","family":"Spector","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Juan F.","family":"Santana","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Arya","family":"Zandvakili","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"David 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A"},{"issue":"6122","key":"ppat.1013374.ref006","doi-asserted-by":"crossref","first-page":"950","DOI":"10.1126\/science.1229386","article-title":"Precise maps of RNA polymerase reveal how promoters direct initiation and pausing","volume":"339","author":"H Kwak","year":"2013","journal-title":"Science"},{"issue":"12","key":"ppat.1013374.ref007","doi-asserted-by":"crossref","first-page":"1198","DOI":"10.1038\/nmeth.4435","article-title":"Thiol-linked alkylation of RNA to assess expression dynamics","volume":"14","author":"VA Herzog","year":"2017","journal-title":"Nat Methods"},{"issue":"7765","key":"ppat.1013374.ref008","doi-asserted-by":"crossref","first-page":"419","DOI":"10.1038\/s41586-019-1369-y","article-title":"scSLAM-seq reveals core features of transcription dynamics in single cells","volume":"571","author":"F Erhard","year":"2019","journal-title":"Nature"},{"issue":"1","key":"ppat.1013374.ref009","doi-asserted-by":"crossref","DOI":"10.1128\/mBio.02047-18","article-title":"Nucleotide resolution comparison of transcription of human cytomegalovirus and host genomes reveals universal use of RNA polymerase II elongation control driven by dissimilar core promoter elements","volume":"10","author":"M Parida","year":"2019","journal-title":"mBio"},{"issue":"8","key":"ppat.1013374.ref010","doi-asserted-by":"crossref","DOI":"10.1128\/JVI.02184-17","article-title":"Herpes simplex virus 1 dramatically alters loading and positioning of RNA polymerase II on host genes early in infection","volume":"92","author":"CH Birkenheuer","year":"2018","journal-title":"J Virol"},{"issue":"2","key":"ppat.1013374.ref011","article-title":"Reactivation of epstein-barr virus from latency involves increased RNA polymerase activity at CTCF binding sites on the viral genome","volume":"97","author":"LEM Dunn","year":"2023","journal-title":"J Virol"},{"issue":"5","key":"ppat.1013374.ref012","doi-asserted-by":"crossref","DOI":"10.1371\/journal.ppat.1010992","article-title":"Decoding murine cytomegalovirus","volume":"19","author":"M Lodha","year":"2023","journal-title":"PLoS Pathog"},{"issue":"5","key":"ppat.1013374.ref013","doi-asserted-by":"crossref","DOI":"10.1128\/jvi.00381-23","article-title":"HSV-1 infection induces a downstream shift of promoter-proximal pausing for host genes","volume":"97","author":"E Wei\u00df","year":"2023","journal-title":"J Virol"},{"issue":"4","key":"ppat.1013374.ref014","article-title":"Human cytomegalovirus IE2 drives transcription initiation from a select subset of late infection viral promoters by host RNA polymerase II","volume":"16","author":"M Li","year":"2020","journal-title":"PLoS Pathog"},{"issue":"8","key":"ppat.1013374.ref015","article-title":"Cytomegalovirus late transcription factor target sequence diversity orchestrates viral early to late 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Spector","year":"2024","journal-title":"Nucleic Acids Res"},{"key":"ppat.1013374.ref022","article-title":"A unified view of the sequence and functional organization of the human RNA polymerase II promoter","author":"DS Luse","year":"2020","journal-title":"Nucleic Acids Res"},{"issue":"4645","key":"ppat.1013374.ref023","doi-asserted-by":"crossref","first-page":"159","DOI":"10.1126\/science.6322309","article-title":"Cytomegalovirus replicates in differentiated but not in undifferentiated human embryonal carcinoma cells","volume":"224","author":"E G\u00f6ncz\u00f6l","year":"1984","journal-title":"Science"},{"issue":"4","key":"ppat.1013374.ref024","doi-asserted-by":"crossref","first-page":"1581","DOI":"10.1128\/JVI.75.4.1581-1593.2001","article-title":"Reactivation of the human cytomegalovirus major immediate-early regulatory region and viral replication in embryonal NTera2 cells: role of trichostatin A, retinoic acid, and deletion of the 21-base-pair repeats and modulator","volume":"75","author":"JL Meier","year":"2001","journal-title":"J Virol"},{"issue":"24","key":"ppat.1013374.ref025","doi-asserted-by":"crossref","first-page":"12284","DOI":"10.1128\/JVI.00965-15","article-title":"Differentiation-coupled induction of human cytomegalovirus replication by union of the major enhancer retinoic acid, cyclic AMP, and NF-\u03baB response elements","volume":"89","author":"J Yuan","year":"2015","journal-title":"J Virol"},{"issue":"34","key":"ppat.1013374.ref026","doi-asserted-by":"crossref","first-page":"31793","DOI":"10.1074\/jbc.M102306200","article-title":"Flavopiridol inactivates P-TEFb and blocks most RNA polymerase II transcription in vivo","volume":"276","author":"SH Chao","year":"2001","journal-title":"J Biol Chem"},{"issue":"17","key":"ppat.1013374.ref027","doi-asserted-by":"crossref","first-page":"8495","DOI":"10.1128\/JVI.00416-10","article-title":"Phorbol ester-induced human cytomegalovirus major immediate-early (MIE) enhancer activation 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products.","volume-title":"Cytomegaloviruses: Molecular Biology and Immunology","author":"JL Meier","year":"2006"},{"issue":"1","key":"ppat.1013374.ref034","doi-asserted-by":"crossref","DOI":"10.1128\/spectrum.03144-22","article-title":"Critical role for the human cytomegalovirus major immediate early proteins in recruitment of RNA polymerase II and H3K27Ac to an enhancer-like element in oriLyt","volume":"11","author":"E Forte","year":"2023","journal-title":"Microbiol Spectr"},{"issue":"11","key":"ppat.1013374.ref035","doi-asserted-by":"crossref","DOI":"10.1128\/JVI.00183-21","article-title":"Epigenetic reprogramming of host and viral genes by Human Cytomegalovirus infection in Kasumi-3 myeloid progenitor cells at early times post-infection","volume":"95","author":"E Forte","year":"2021","journal-title":"J Virol"},{"issue":"20","key":"ppat.1013374.ref036","doi-asserted-by":"crossref","first-page":"9998","DOI":"10.1128\/JVI.01297-06","article-title":"Autorepression of the human cytomegalovirus major immediate-early promoter\/enhancer at late times of infection is mediated by the recruitment of chromatin remodeling enzymes by IE86","volume":"80","author":"M Reeves","year":"2006","journal-title":"J Virol"},{"issue":"4","key":"ppat.1013374.ref037","article-title":"Chromatin control of human cytomegalovirus infection","volume":"14","author":"SM Matthews","year":"2023","journal-title":"mBio"},{"key":"ppat.1013374.ref038","doi-asserted-by":"crossref","first-page":"476","DOI":"10.3389\/fcimb.2020.00476","article-title":"Control of immediate early gene expression for human cytomegalovirus reactivation","volume":"10","author":"D Collins-McMillen","year":"2020","journal-title":"Front Cell Infect Microbiol"},{"issue":"22","key":"ppat.1013374.ref039","doi-asserted-by":"crossref","first-page":"11167","DOI":"10.1128\/JVI.01218-08","article-title":"Temporal dynamics of cytomegalovirus chromatin assembly in productively infected human cells","volume":"82","author":"A Nitzsche","year":"2008","journal-title":"J Virol"},{"issue":"18","key":"ppat.1013374.ref040","doi-asserted-by":"crossref","first-page":"9817","DOI":"10.1128\/JVI.00581-12","article-title":"Histone H3 lysine 4 methylation marks postreplicative human cytomegalovirus chromatin","volume":"86","author":"A Nitzsche","year":"2012","journal-title":"J Virol"},{"issue":"2","key":"ppat.1013374.ref041","doi-asserted-by":"crossref","DOI":"10.1128\/Spectrum.00539-21","article-title":"Characteristics of immediate-Early 2 (IE2) and UL84 proteins in UL84-independent strains of human cytomegalovirus (HCMV)","volume":"9","author":"S Manska","year":"2021","journal-title":"Microbiol Spectr"},{"issue":"13","key":"ppat.1013374.ref042","doi-asserted-by":"crossref","first-page":"6391","DOI":"10.1128\/JVI.00061-09","article-title":"Breaking human cytomegalovirus major immediate-early gene silence by vasoactive intestinal peptide stimulation of the protein kinase A-CREB-TORC2 signaling cascade in human pluripotent embryonal NTera2 cells","volume":"83","author":"J Yuan","year":"2009","journal-title":"J Virol"},{"issue":"2","key":"ppat.1013374.ref043","doi-asserted-by":"crossref","first-page":"1246","DOI":"10.1128\/jvi.71.2.1246-1255.1997","article-title":"Effect of a modulator deletion on transcription of the human cytomegalovirus major immediate-early genes in infected undifferentiated and differentiated cells","volume":"71","author":"JL Meier","year":"1997","journal-title":"J Virol"},{"issue":"4","key":"ppat.1013374.ref044","doi-asserted-by":"crossref","first-page":"1870","DOI":"10.1128\/JVI.75.4.1870-1878.2001","article-title":"Human cytomegalovirus with IE-2 (UL122) deleted fails to express early lytic genes","volume":"75","author":"A Marchini","year":"2001","journal-title":"J Virol"},{"issue":"4","key":"ppat.1013374.ref045","doi-asserted-by":"crossref","DOI":"10.1093\/nar\/gni035","article-title":"Simple and highly efficient BAC recombineering using galK selection","volume":"33","author":"S Warming","year":"2005","journal-title":"Nucleic Acids Res"},{"issue":"19","key":"ppat.1013374.ref046","doi-asserted-by":"crossref","first-page":"11088","DOI":"10.1093\/nar\/gkx724","article-title":"Oxidative stress rapidly stabilizes promoter-proximal paused Pol II across the human genome","volume":"45","author":"KA Nilson","year":"2017","journal-title":"Nucleic Acids Res"},{"issue":"3","key":"ppat.1013374.ref047","doi-asserted-by":"crossref","DOI":"10.1186\/gb-2009-10-3-r25","article-title":"Ultrafast and memory-efficient alignment of short DNA sequences to the human genome","volume":"10","author":"B Langmead","year":"2009","journal-title":"Genome Biol"},{"issue":"6","key":"ppat.1013374.ref048","doi-asserted-by":"crossref","first-page":"841","DOI":"10.1093\/bioinformatics\/btq033","article-title":"BEDTools: a flexible suite of utilities for comparing genomic features","volume":"26","author":"AR 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