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Quantitative proteomic discovery of dynamic epigenome changes that control human cytomegalovirus (HCMV) infection.

ABSTRACT: This work represents the first comprehensive quantitative analysis of global histone post-translational modifications (PTMs) from a virus infection, namely human cytomegalovirus (HCMV) infection. We used a nanoLC-MS/MS platform to identify and quantify the dynamic histone H3 and H4 PTMs expressed during HCMV replication in primary fibroblasts. Specifically, we examined the changes in histone PTMs over a 96 h time course to sample the immediate early (IE), early (E), and late (L) stages of viral infection. Several changes in histone H3 and H4 PTMs were observed, including a marked increase in H3K79me2 and H3K27me3K36me2, and a decrease in H4K16ac, highlighting likely epigenetic strategies of transcriptional activation and silencing during HCMV lytic infection. Heavy methyl-SILAC (hm-SILAC) was used to further confirm the histone methylation flux (especially for H3K79) during HCMV infection. We evaluated DOT1L (the H3K79 methyltransferase) mRNA levels in mock and HCMV-infected cells over a 96 h time course, and observed a significant increase in this methyltransferase as early as 24 hpi showing that viral infection up-regulates DOT1L expression, which drives H3K79me2. We then used shRNA to create a DOT1L knockdown cell population, and found that HCMV infection of the knockdown cells resulted in a 10-fold growth defect when compared with infected control cells not subjected to knockdown. This work documents multiple histone PTMs that occur in response to HCMV infection of fibroblasts, and provides a framework for evaluation of the role of epigenetic modifications in the virus-host interaction.

SUBMITTER: O'Connor CM

PROVIDER: S-EPMC4159657 | biostudies-literature | 2014 Sep

REPOSITORIES: biostudies-literature

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Quantitative proteomic discovery of dynamic epigenome changes that control human cytomegalovirus (HCMV) infection.

O'Connor Christine M CM DiMaggio Peter A PA Shenk Thomas T Garcia Benjamin A BA

Molecular & cellular proteomics : MCP 20140701 9

This work represents the first comprehensive quantitative analysis of global histone post-translational modifications (PTMs) from a virus infection, namely human cytomegalovirus (HCMV) infection. We used a nanoLC-MS/MS platform to identify and quantify the dynamic histone H3 and H4 PTMs expressed during HCMV replication in primary fibroblasts. Specifically, we examined the changes in histone PTMs over a 96 h time course to sample the immediate early (IE), early (E), and late (L) stages of viral ...[more]

PMID: 24987098

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Project description:Individuals lacking functional natural killer (NK) cells suffer severe, recurrent infections with cytomegalovirus (CMV), highlighting the critical role of NK cells in antiviral defense. Therefore, ongoing attempts to develop an efficacious vaccine to prevent CMV infection should potentially aim to elicit NK-cell antiviral responses as an accessory to conventional T- and B-cell based approaches. In this regard, CMV infection provokes marked phenotypic and functional differentiation of the NK-cell compartment, including development of adaptive NK cells that exhibit enhanced antiviral activity. We examined longitudinal blood samples collected from 40 CMV-seronegative adolescents to ascertain whether a CMV glycoprotein B (gB) vaccine in the absence of CMV infection can stimulate differentiation or expansion of CMV-associated subsets of NK cells. Study participants uniformly lacked the CMV-dependent NKG2C+ subset of NK cells, suggesting that an adjuvanted CMV gB vaccine alone is an inadequate stimulus for sustained expansion of these cells. In contrast, we observed unexpected dynamic fluctuations in the frequency of NK cells lacking FcRγ, EAT-2, and SYK, which were independent of vaccination or CMV infection. Whereas, FcRγneg NK cells in CMV infection are reported to express increased levels of the maturation marker CD57, the FcRγneg NK cells observed in our CMV-negative vaccine cohort express less CD57 than their FcRγ+ counterparts. The FcRγneg NK cells in CMV-negative individuals were also functionally distinct from this subset in CMV infection, exhibiting comparable IFN-γ production and degranulation as FcRγ+ NK cells in response to cytokine or antibody-dependent stimuli. These results suggest that frequencies of some NK cell subsets may increase in response to unknown environmental or inflammatory cues distinct from that which occurs after CMV infection. Greater understanding of the nature of the signals driving CMV-independent accumulation of these subsets should permit development of mechanisms to facilitate vaccine-driven expansion of CMV-reactive NK cells.

Dataset Information

Quantitative proteomic discovery of dynamic epigenome changes that control human cytomegalovirus (HCMV) infection.

Publications

Quantitative proteomic discovery of dynamic epigenome changes that control human cytomegalovirus (HCMV) infection.

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