Dissecting HSV-1-induced host shut-off at RNA level

Abstract

26Herpes simplex virus 1 (HSV-1) installs a profound host shut-off during lytic infection. 27 The virion host shut-off (vhs) protein plays a key role in this process by efficiently 28 cleaving both host and viral mRNAs in a translation-initiation-dependent manner. 29 Furthermore, the onset of viral DNA replication is accompanied by a rapid decline in 30 transcriptional activity of the host genome. Both mechanisms have tremendous impact 31 on the RNA expression profile of the infected cells. To dissect their relative 32 contributions and elucidate gene-specific host transcriptional responses throughout 33 the first 8h of lytic HSV-1 infection, we here employed RNA-seq of total, newly 34 transcribed (4sU-labelled) and chromatin-associated RNA in wild-type (WT) and vhs 35 infection of primary human fibroblasts. Following virus entry, vhs activity rapidly 36 plateaued at an elimination rate of around 30% of cellular mRNAs per hour until 8h p.i. 37In parallel, host transcriptional activity dropped down to 10-20%. While the combined 38 effects of both phenomena dominated infection-induced changes in total RNA, 39 extensive gene-specific transcriptional regulation was observable in chromatin-40 associated RNA. This was surprisingly concordant between WT HSV-1 and its vhs 41 mutant and at least in parts mediated by the embryonic transcription factor DUX4. 42 Furthermore, both WT and vhs infection induced strong transcriptional up-regulation 43 of a small subset of genes. Most of these were either poorly or not at all expressed 44 prior to infection but already primed by H3K4me3 histone marks at their promoters. 45 Most interestingly, analysis of chromatin-associated RNA revealed vhs-nuclease- 46 activity-dependent transcriptional down-regulation of at least 150 cellular genes, in 47 particular of many genes encoding integrin adhesome and extracellular matrix 48 components. This was accompanied by a vhs-dependent reduction in protein levels by 49 8h p.i. for many of these genes. In summary, our study provides a comprehensive 50 picture of the molecular mechanisms that govern cellular RNA metabolism during the 51 first 8h of lytic HSV-1 infection. 52 53 4 Author Summary 54The HSV-1 virion host shut-off (vhs) protein efficiently cleaves both host and viral 55 mRNAs in a translation-dependent manner. In this study, we model and quantify 56 changes in vhs activity as well as the virus-induced global loss of host transcriptional 57 activity during productive HSV-1 infection. In general, HSV-1-induced alterations in 58 total RNA levels were found to be predominantly shaped by these two global processes 59 rather than gene-specific regulation. In contrast, chromatin-associated RNA depicted 60 gene-specific transcriptional changes. This revealed highly concordant transcriptional 61 changes in WT and vhs infection, confirmed DUX4 as a key transcriptional regulator 62 in HSV-1 infection and depicted vhs-dependent, transcriptional down-regulation of the 63 integrin adhesome and extracellular matrix. The latt...

HSV-1 infection induces a downstream shift of the +1 nucleosome

“…Notably, we previously showed that transcriptional activity on host genes drops substantially in the first 4 h of infection to only 40% of transcription in uninfected cells, which was further halved until 8 h p.i. (19). Thus, the onset of the extension of accessible chromatin regions at promoters into downstream (for pattern (i)) and upstream (for pattern (ii)) regions follows the onset of the drop in host transcription during infection.…”

“…To correlate transcriptional activity of genes to the observed patterns in chromatin accessibility changes, we analyzed RNA-seq data of chromatin-associated RNA for mock and 8 h p.i. WT HSV-1 infection from our recent study (20). Chromatin-associated RNA depicts ongoing nascent transcription on the chromatin.…”

“…For this purpose, we analyzed RNA-seq data of chromatin-associated RNA, which depicts nascent transcription, for mock and 8 h p.i. WT HSV-1 infection from our recent study (20). We also included in this analysis the 2,668 genes (denoted as NA group) without significant chromatin accessibility changes that were excluded from Fig 1a . Interestingly, metagene analyses for these genes also revealed a slight broadening of the TSS peak into downstream regions, although this was much less pronounced than for pattern (i) clusters and therefore not detected by RegCFinder (Sup.…”

“…Chromatin-associated RNA depicts ongoing nascent transcription on the chromatin. Furthermore, changes in chromatinassociated RNA better reflect transcriptional changes in HSV-1 infection than total RNA measurements as the latter are confounded by vhs-mediated mRNA degradation (20). Our analysis also included the 2,668 genes (denoted as NA group) without significant chromatin accessibility changes that were excluded from Fig.…”

“…vhs is a nuclease delivered by the tegument of the incoming viral particles that cleaves both viral and host mRNAs (17)(18)(19). We recently demonstrated that vhs continuously degrades about 30 % of host mRNAs per hour during the first 8 h of high multiplicity lytic infection of primary human fibroblasts (20).…”