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Human cytomegalovirus IE2 drives transcription initiation from a select subset of late infection viral promoters by host RNA polymerase II.

ABSTRACT: Herpesvirus late promoters activate gene expression after viral DNA synthesis has begun. Alphaherpesviruses utilize a viral immediate-early protein to do this, whereas beta- and gammaherpesviruses primarily use a 6-member set of viral late-acting transcription factors (LTF) that are drawn to a TATT sequence in the late promoter. The betaherpesvirus, human cytomegalovirus (HCMV), produces three immediate-early 2 protein isoforms, IE2-86, IE2-60, IE2-40, late in infection, but whether they activate late viral promoters is unknown. Here, we quickly degrade the IE2 proteins in late infection using dTag methodology and analyze effects on transcription using customized PRO-Seq and computational methods combined with multiple validation methods. We discover that the IE2 proteins selectively drive RNA Pol II transcription initiation at a subset of viral early-late and late promoters common to different HCMV strains, but do not substantially affect Pol II transcription of the 9,942 expressed host genes. Most of the IE2-activated viral late infection promoters lack the TATT sequence bound by the HCMV UL87-encoded LTF. The HCMV TATT-binding protein is not mechanistically involved in late RNA expression from the IE2-activated TATT-less UL83 (pp65) promoter, as it is for the TATT-containing UL82 (pp71) promoter. While antecedent viral DNA synthesis is necessary for transcription from the late infection viral promoters, continued viral DNA synthesis is unnecessary. We conclude that in late infection the IE2 proteins target a distinct subset of HCMV early-late and late promoters for transcription initiation by RNA Pol II. Commencement of viral DNA replication renders the HCMV genome late promoters susceptible to late-acting viral transcription factors.

SUBMITTER: Li M

PROVIDER: S-EPMC7162547 | biostudies-literature | 2020 Apr

REPOSITORIES: biostudies-literature

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Human cytomegalovirus IE2 drives transcription initiation from a select subset of late infection viral promoters by host RNA polymerase II.

Li Ming M Ball Christopher B CB Collins Geoffrey G Hu Qiaolin Q Luse Donal S DS Price David H DH Meier Jeffery L JL

PLoS pathogens 20200406 4

Herpesvirus late promoters activate gene expression after viral DNA synthesis has begun. Alphaherpesviruses utilize a viral immediate-early protein to do this, whereas beta- and gammaherpesviruses primarily use a 6-member set of viral late-acting transcription factors (LTF) that are drawn to a TATT sequence in the late promoter. The betaherpesvirus, human cytomegalovirus (HCMV), produces three immediate-early 2 protein isoforms, IE2-86, IE2-60, IE2-40, late in infection, but whether they activat ...[more]

PMID: 32251483

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Project description:Human cytomegalovirus (HCMV) immediate-early 2 (IE2) protein is a multifunctional transcription factor that is essential for lytic HCMV infection. IE2 functions as an activator of viral early genes, negatively regulates its own promoter, and is required for viral replication. The mechanisms by which IE2 executes these distinct functions are incompletely understood. Using PRO-Seq, which profiles nascent transcripts, and a recently developed DFF-chromatin immunoprecipitation (DFF-ChIP; employs chromatin digestion by the endonuclease DNA fragmentation factor prior to IP) approach that resolves occupancy and local chromatin environment, we show that IE2 controls viral gene transcription in three distinct capacities during late HCMV infection and reveal mechanisms that involve direct binding of IE2 to viral DNA. IE2 represses a subset of viral promoters by binding within their core promoter regions and blocking the assembly of preinitiation complexes (PICs). Remarkably, IE2 forms a repressive complex at the major immediate-early promoter region involving direct association of IE2 with nucleosomes and TBP. IE2 stimulates transcription by binding nearby, but not within, core promoter regions. In addition, IE2 functions as a direct roadblock to transcription elongation. At one locus, this function of IE2 appears to be important for the synthesis of a spliced viral RNA. Consistent with the minimal observed effects of IE2 depletion on host gene transcription, IE2 does not functionally engage the host genome. Our results reveal mechanisms of transcriptional control by IE2, uncover a previously unknown function of IE2 as a Pol II elongation modulator, and demonstrate that DFF-ChIP is a useful tool for probing transcription factor occupancy and interactions between transcription factors and nucleosomes at high resolution. IMPORTANCE HCMV infects more than half of the world population and persists lifelong in its hosts. Although generally asymptomatic, HCMV infection can lead to life-threating disease in immunosuppressed individuals. Moreover, HCMV is the leading infectious cause of birth defects in the United States. As there are no vaccines effective against HCMV and antiviral drugs exhibit toxicity and are undermined by resistant HCMV variants, other vulnerabilities in HCMV must be explored. Here, we characterize the mechanism by which IE2 controls transcription during late HCMV infection. We demonstrate that IE2 engages numerous consensus sites across the HCMV genome and functions as an activator, repressor, or elongation modulator depending on the context of IE2 binding sites in relation to Pol II initiation and elongation complexes. Our findings have important implications for the ongoing exploration of IE2 as an antiviral drug target.

Dataset Information

Human cytomegalovirus IE2 drives transcription initiation from a select subset of late infection viral promoters by host RNA polymerase II.

Publications

Human cytomegalovirus IE2 drives transcription initiation from a select subset of late infection viral promoters by host RNA polymerase II.

Similar Datasets

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