Biology subjects

Doelken, L.

Publications and source records attributed to Doelken, L..

15 recordsLinked to original sources

bioRxiv2026

Herpes simplex virus infection promotes ALS pathology through ICP0-mediated PML body disruption

Transactive response DNA binding protein 43 kDa (TDP-43) pathology, is a central molecular hallmark of amyotrophic lateral sclerosis (ALS). However, the underlying triggers are incompletely understood. Here, we show that infection with herpes simplex virus (HSV) induces molecular hallmarks of ALS in various in vitro and in vivo models and is associated with an increased risk of ALS in human population data. German healthcare provider data (n = 238,440) and herpesvirus serology of an ALS patient and control cohort (n = 1,100) showed that HSV infection elevated the ALS risk by 210% and odds by [~]65%, respectively. On a molecular level, HSV infection promoted TDP-43 pathology in neuronal cell models, human iPSC-derived motoneurons and cerebral organoids, mice, and human tissue sections. This effect was triggered by HSV-1 or 2, but not by several other related herpesviruses. Mechanistically, the infected cell protein 0 (ICP0) of HSV-1/2 drives TDP-43 pathology by disturbance of promyelocytic leukemia nuclear bodies (PML-NBs), thereby abrogating TDP-43 SUMO2/3ylation. Taken together, we reveal a previously unrecognized association between HSV infection and ALS and clarify the underlying molecular mechanism that drives TDP-43 pathology. Our data may guide future studies into therapeutic and prophylactic interventions against ALS.

neuroscience↗

bioRxiv2025

Unveiling immune interference: How the dendritic cell response to co-infection with Aspergillus fumigatus is modulated by human cytomegalovirus and its virokine CMVIL-10

Human cytomegalovirus (HCMV) is a master of immune evasion and a potent modulator of the human immune system. The best-characterized mechanism employed by HCMV to suppress host immunity is the production of a viral interleukin-10 homolog (CMVIL-10). While CMVIL-10 is known to suppress immune responses and promote viral persistence, its capacity to promote increased susceptibility to co-infecting pathogens like Aspergillus fumigatus remains unknown. Therefore, we studied the impact of wild-type (WT) HCMV (strain TB40 BAC4), a CMVIL-10-deficient HCMV mutant ({Delta}UL111A), and recombinant CMVIL-10 on the immune activity of monocyte-derived dendritic cells (moDCs) during co-infection with A. fumigatus. Using a combination of transcriptomic and phenotypic readouts, our data revealed a strong and time-dependent immuno-paralytic effect of HCMV by suppressing pathogen recognition pathways, cytokine production, DC maturation, and expression of genes that are essential for host defense and tissue repair. Although infection with {Delta}UL111A lacking CMVIL-10 led to stronger expression of type I interferons, IFN-{gamma}-inducible chemokines, and proinflammatory cytokines than WT infection, interference with antifungal immune defense and fungal clearance during co-infection was largely similar between both strains. The limited effect of CMVIL-10 on antifungal immune defense persisted even after prolonged pre-exposure of DCs to the recombinant virokine. In summary, although CMVIL-10 contributes to shaping an anti-inflammatory environment, HCMVs suppression of antifungal immunity appears to be multifactorial, with CMVIL-10 alone playing a rather subtle role in altering DC responses to A. fumigatus during viral-fungal co-infection. ImportanceHuman cytomegalovirus (HCMV) is a highly prevalent herpesvirus that establishes lifelong latency and frequently reactivates in immunocompromised individuals, including hematopoietic stem cell transplant recipients. Reactivation not only causes direct disease but also increases the risk of secondary infections, such as invasive pulmonary aspergillosis caused by Aspergillus fumigatus. Specifically, studies estimated that about 6-25% of critically ill HCMV-positive patients develop HCMV-associated pulmonary aspergillosis. However, the mechanisms by which HCMV creates a permissive environment for fungal superinfection remain poorly understood. HCMV encodes a viral homolog of interleukin-10 (CMVIL-10), which mimics host IL-10 and elicits potent immunomodulatory activity. Here, we show that CMVIL-10 dampens specific anti-viral responses, DC activation, and cytokine signaling. However, HCMV-mediated impairment of fungal control in co-infection settings occurred largely independent of CMVIL-10 expression. These findings suggest that HCMV undermines antifungal defenses through multifactorial mechanisms beyond CMVIL-10, highlighting the need for targeted strategies to restore immune function in high-risk patients.

immunology↗

bioRxiv2024

Identifying Modulators of Cellular Responses by Heterogeneity-sequencing

The response of individual cells to drug treatment, virus infections or other molecular stimuli is highly heterogeneous and depends on the cells initial state. Library preparation for single-cell transcriptomics is destructive, precluding a direct comparison between the initial state and the stimulus outcome. Consequently, current methods are restricted to identifying correlative associations rather than resolving causal drivers of heterogeneous outcomes. We developed Heterogeneity-seq, which combines single-cell RNA-seq with metabolic RNA labeling (scSLAM-seq) and double machine learning to overcome this limitation. By leveraging simultaneous measurements of unlabeled and labeled RNA in individual cells, Heterogeneity-seq uncovers the transition from pre-stimulated cell states to distinct stimulation outcomes across thousands of cells. These links enable the identification of factors that causally govern heterogeneous cellular responses. We used Heterogeneity-seq to identify both known and novel genes that drive responses to drug treatment, as well as pro- and antiviral host factors governing cytomegalovirus infection.

bioinformatics↗

bioRxiv2024

HSV-1 infection induces a downstream-shift of +1 nucleosomes

Herpes simplex virus 1 (HSV-1) infection induces a loss of host transcriptional activity and widespread disruption of host transcription termination, which leads to an induction of open chromatin downstream of genes. In this study, we show that lytic HSV-1 infection also leads to an extension of chromatin accessibility at promoters into downstream regions. This is most prominent for highly expressed genes and independent of the immediate-early proteins ICP0, ICP22, and ICP27 and the virion host shutoff protein vhs. ChIPmentation of the noncanonical histone variant H2A.Z, which is strongly enriched at +1 and -1 nucleosomes, indicated that these chromatin accessibility changes are linked to a downstream shift of +1 nucleosomes. In yeast, downstream shifts of +1 nucleosomes are induced by RNA Polymerase II (Pol II) degradation. Accordingly, irreversible depletion of Pol II from genes in human cells using -amanitin altered +1 nucleosome positioning similar to lytic HSV-1 infection. Consequently, treatment with phosphonoacetic acid (PAA) and knockout of ICP4, which both prevent viral DNA replication and alleviate the loss of Pol II from host genes, largely abolished the downstream extension of accessible chromatin in HSV-1 infection. In the absence of viral DNA replication, doxycycline-induced expression of ICP27, which redirects Pol II from gene bodies into intergenic regions by disrupting transcription termination, induced an attenuated effect that was further enhanced by co-expression of ICP22. In summary, our study provides strong evidence that HSV-1-induced depletion of Pol II from the host genome leads to a downstream shift of +1 nucleosomes at host promoters. ImportanceLytic herpes simplex virus 1 (HSV-1) infection leads to a profound host transcription shutoff. Loss of RNA Polymerase II (Pol II) in yeast has previously been shown to relax +1 nucleosome positioning to more thermodynamically favorable sites downstream of transcription start sites. Here, we show that a similar phenomenon is likely at play in lytic HSV-1 infection. Sequencing of accessible chromatin revealed a widening of nucleosome-free regions at host promoters into downstream regions. By mapping genome-wide positions of the noncanonical histone variant H2A.Z enriched at +1 and -1 nucleosomes, we demonstrate a downstream shift of +1 nucleosomes for most cellular genes in lytic HSV-1 infection. As chemical depletion of Pol II from genes also leads to a downstream shift of +1 nucleosomes in human cells, changes in chromatin architecture at promoters in HSV-1 infection are likely a consequence of HSV-1-induced loss of Pol II activity from the host genome.

bioinformatics↗

bioRxiv2024

Building Blocks of Understanding: Constructing a Reverse Genetics Platform for studying determinants of SARS-CoV-2 replication.

To better understand viral pathogenesis, host-virus interactions, and potential therapeutic interventions, the development of robust reverse genetics systems for SARS-CoV-2 is crucial. Here, we present a reverse genetics platform that enables the efficient manipulation, assembly, and rescue of recombinant SARS-CoV-2. The versatility of our reverse genetics system was demonstrated by generating recombinant SARS-CoV-2 viruses. We used this system to generate N501Y and Y453F spike protein mutants. Characterization studies revealed distinct phenotypic effects, impact on viral fitness, cell binding, and replication kinetics. We also investigated a recently discovered priming site for NSP9, which is postulated to produce a short RNA antisense leader sequence. By introducing the U76G mutation into the 5UTR, we show that this priming site is necessary for the correct production of genomic and subgenomic RNAs, and also for efficient viral replication. In conclusion, our developed reverse genetics system provides a robust and adaptable platform for the efficient generation of recombinant SARS-CoV-2 viruses for their comprehensive characterization. Significance statementIn this study, we present a versatile reverse genetics platform facilitating the efficient manipulation, assembly, and rescue of recombinant SARS-CoV-2. Demonstrating its adaptability, we successfully engineered N501Y and Y453F spike protein mutants, each exhibiting distinct phenotypic effects on viral fitness, cell binding, and replication kinetics. We also investigated a novel negative sense priming site for NSP9, demonstrating a role in RNA production and viral replication. This straightforward reverse genetic system is therefore a powerful tool to generate recombinant viruses for advancing our understanding of SARS-CoV-2 biology.

microbiology↗

bioRxiv2023

mRNA N1-2'O-methylation by CMTR1 affects NVL2 mRNA splicing

Cap0-mRNA is characterized by a 5-5triphosphate-linked N7-methylated guanosine(m7G). In higher eukaryotes, the methyltransferase CMTR1 additionally methylates the 2O-position of the penultimate mRNA nucleotide(N1) ribose (cap1-mRNA). While the m7G cap is essential for mRNA export and translation initiation by the eIF4F complex, the N1-2O-methylation prevents recognition of cap1-mRNA by the antiviral RNA receptors RIG-I and IFIT1, but a function beyond immunotolerance remained elusive. Here, we generated CMTR1-knockout(CMTR1-/-) cells and found that type-I-interferon(IFN-I) treatment resulted in IFIT1-mediated reduction of cell viability and broad mRNA translation. Consequently, stimulation of the antiviral receptor RIG-I in CMTR1-/- cells revealed an IFIT1 dependent dramatic reduction of IFN-I and chemokine protein induction, demonstrating the importance of N1-2O-methylation for antiviral responses. Additionally, IFN-I- and IFIT1-independent effects were observed: CMTR1-/- cells were smaller, divided slower, and exhibited a reduced transcription of mRNAs coding ribosomal proteins (RP), 5TOP-RNA and snoRNA host genes(SNHG). Additionally, proteome and transcriptome analysis revealed that expression of NVL2, an essential factor in ribosome biogenesis, is strongly suppressed by an alternative-splicing event of NVL2 mRNA in CMTR1-/- cells. This reduction could only be rescued by catalytically active CMTR1. Altogether, besides antiviral immunity N1-2O-methylation by CMTR1 has broad effects on cellular physiology and controls splicing of NVL2.

molecular biology↗

bioRxiv2023

The Cytomegalovirus M35 Protein Modulates Transcription of Ifnb1 and Other IRF3-Driven Genes by Direct Promoter Binding

Induction of type I interferon (IFN) gene expression is among the first lines of cellular defence a virus encounters during primary infection. We previously identified the tegument protein M35 of murine cytomegalovirus (MCMV) as an essential antagonist of this antiviral system. M35 localizes to the nucleus and interferes with type I IFN induction downstream of pattern-recognition receptor (PRR) activation. Here, we report structural and mechanistic details of M35s function. Using electrophoretic mobility shift assays (EMSA), we demonstrate that purified M35 protein specifically binds to the regulatory DNA element that governs transcription of the first type I IFN gene induced in non-immune cells, Ifnb1. Determination of M35s crystal structure combined with reverse genetics revealed that homodimerisation is a key feature for M35s immunomodulatory activity. DNA-binding sites of M35 overlapped with the recognition elements of interferon regulatory factor 3 (IRF3), a key transcription factor activated by PRR signalling. Chromatin immunoprecipitation (ChIP) showed reduced binding of IRF3 to the host Ifnb1 promoter in the presence of M35. We furthermore defined the IRF3-dependent and the type I IFN signalling-responsive genes in murine fibroblasts by RNA sequencing of metabolically labelled transcripts (SLAM-seq), and assessed M35s global effect on gene expression. Stable expression of M35 broadly influenced the transcriptome in untreated cells and specifically down-regulated basal expression of IRF3-dependent genes, and during MCMV infection, M35 impaired expression of IRF3-responsive genes aside of Ifnb1. Our results suggest that M35-DNA binding directly antagonises gene induction by IRF3 and impairs the antiviral response more broadly than formerly recognised. ImportanceReplication of the ubiquitous human cytomegalovirus (CMV) in healthy individuals mostly goes unnoticed, but can impair foetal development or cause life-threatening symptoms in immunosuppressed or -deficient patients. Like other herpesviruses, CMV extensively manipulates its hosts and establishes lifelong latent infections. Murine CMV (MCMV) presents an important model system as it allows the study of CMV infection in the host organism. We previously showed that during entry, MCMV virions release the evolutionary conserved protein M35 protein to immediately dampen the antiviral type I interferon (IFN) response induced by pathogen detection. Here we show that M35 dimers bind to regulatory DNA elements and interfere with recruitment of interferon regulatory factor 3 (IRF3), a key factor for antiviral gene expression. Thereby, M35 interferes with expression of type I IFNs and other IRF3-dependent genes. Unrelated proteins from other herpesviruses employ the same mechanism, reflecting the importance for herpesviruses to avoid IRF3-mediated gene induction.

molecular biology↗

bioRxiv2022

Decoding murine cytomegalovirus

The genomes of both human cytomegalovirus (HCMV) and murine cytomegalovirus (MCMV) were first sequenced over 20 years ago. Similar to HCMV, the MCMV genome had initially been proposed to harbor {approx}170 open reading frames (ORFs). More recently, omics approaches revealed HCMV gene expression to be substantially more complex comprising several hundreds of translated ORFs. Here, we provide a state-of-the art reannotation of lytic MCMV gene expression based on integrative analysis of a large set of omics data. Our data reveal 363 viral transcription start sites (TiSS) that give rise to 380 and 454 viral transcripts and ORFs, respectively. The latter include >200 small ORFs, some of which represented the most highly expressed viral gene products. By combining TiSS profiling with metabolic RNA labelling and chemical nucleotide conversion sequencing (dSLAM-seq), we provide a detailed picture of the expression kinetics of viral transcription. This not only resulted in the identification of a novel MCMV immediate early transcript encoding the m166.5 ORF, which we termed ie4, but also revealed a group of well-expressed viral transcripts that are induced later than canonical true late genes and contain an initiator element (Inr) but no TATA- or TATT-box in their core promoters. We show that viral uORFs tune gene expression of longer viral ORFs expressed in cis at translational level. Finally, we identify a truncated isoform of the viral NK-cell immune evasin m145 arising from a viral TiSS downstream of the canonical m145 mRNA. Despite being {approx}5-fold more abundantly expressed than the canonical m145 protein it was not required for downregulating the NK cell ligand, MULT-I. In summary, our work will pave the way for future mechanistic studies on previously unknown cytomegalovirus gene products in an important virus animal model. Author summaryWe conducted a comprehensive characterization and reannotation of murine cytomegalovirus (MCMV) gene expression during lytic infection in murine fibroblasts using an integrative multi-omics approach. This unveiled hundreds of novel transcripts that explained the expression of close to 300 so far unknown viral open reading frames (ORFs). Interestingly, small viral ORFs (sORFs) were amongst the most highly expressed viral gene products and thus presumably encode for important viral microproteins of unknown function. However, we also show that sORFs located upstream of larger ORFs tune the expression of the downstream ORFs at the level of translation. We classified viral transcription start sites (TiSS) based on their expression kinetics obtained by a new combination of metabolic RNA labelling with transcription start sites profiling. This not only identified a so far unknown viral immediate-early transcript (ie4, m166.5 RNA) but also revealed a novel class of viral late transcripts that are expressed later than canonical true late genes and lack TATA box-like motifs. We exemplify for the m145 locus how so far unknown TiSS give rise to abundantly expressed truncated viral proteins. In summary, we provide a state-of-the-art annotation of an important model virus, which will be instrumental for future studies on CMV biology, immunology and pathogenesis.

microbiology↗

bioRxiv2022

HSV-1 infection induces a downstream shift of promoter-proximal pausing linked to H2A.Z-containing +1 nucleosomes

Herpes simplex virus 1 (HSV-1) infection exerts a profound shut-off of host gene expression at multiple levels. Recently, HSV-1 infection was reported to also impact promoter-proximal RNA Polymerase II (Pol II) pausing, a key step in the eukaryotic transcription cycle, with decreased and increased Pol II pausing observed for activated and repressed genes, respectively. Here, we demonstrate that HSV-1 infection induces more complex alterations in promoter-proximal pausing than previously suspected for the vast majority of cellular genes. While pausing is generally retained, it is shifted to more downstream and less well-positioned sites for most host genes. We exclude that this is an artefact of alternative de novo transcription initiation at downstream sites or read-in transcription originating from disruption of transcription termination for upstream genes. Use of downstream 2nd pause sites associated with +1 nucleosomes was previously observed upon NELF depletion. However, downstream shifts of Pol II pausing upon HSV-1 infection are much more pronounced than observed upon NELF depletion. Thus, our study reveals a novel aspect in which HSV-1 infection fundamentally reshapes host transcriptional processes with implications for our understanding of maintenance of promoter-proximal Pol II pausing in eukaryotic cells.

molecular biology↗

bioRxiv2022

U1 snRNP-dependent suppression of miRNA biogenesis by alternative intronic polyadenylation in melanoma

Activation of intronic polyadenylation signals results in premature cleavage and polyadenylation (PCPA). The majority of mammalian miRNAs are also located within intronic regions of protein-coding genes and are transcriptionally co-expressed with their host genes. Here we show that U1-dependent PCPA by telescripting dysregulates miRNA biogenesis. When U1 is reduced, miR-211 levels are decreased as a direct consequence of activation of a newly identified alternative intronic polyadenylation signal located upstream of miR-211 within its host gene TRPM1. Various melanoma cell lines revealed decreased U1 levels and a shift from full-length to truncated TRPM1 isoforms with concomitant decreased miR-211 expression. Modulation of TRPM1 alternative polyadenylation (APA) by morpholino oligonucleotides inhibits and potentially restores miR-211 expression to endogenous levels. This mechanism of intronic PCPA and its effects on miRNA biogenesis represents a previously unrecognized layer of gene expression regulation suitable for therapeutic modulation. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=88 SRC="FIGDIR/small/479622v1_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@169c2f1org.highwire.dtl.DTLVardef@7d8f34org.highwire.dtl.DTLVardef@144b9d8org.highwire.dtl.DTLVardef@10523f1_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2021

HSV-1 and influenza infection induce linear and circular splicing of the long NEAT1 isoform

The herpes simplex virus 1 (HSV-1) virion host shut-off (vhs) protein cleaves both cellular and viral mRNAs by a translation-initiation-dependent mechanism. Here, we show that vhs-mediated degradation of mRNAs leads to an accumulation of circular RNAs (circRNAs) relative to linear mRNAs during HSV-1 infection. Strikingly, we found that circular splicing of the long isoform (NEAT1_2) of the nuclear paraspeckle assembly transcript 1 (NEAT1) was massively induced during HSV-1 infection. In contrast to other circRNAs, induction of the NEAT1_2 circRNA was independent of vhs and occurred while NEAT1_2 was still bound to the chromatin. This was associated with induction of linear splicing of NEAT1_2 both within and downstream of the circRNA. NEAT1_2 splicing was absent in uninfected cells but can be induced by ectopic co-expression of the HSV-1 immediate-early proteins ICP22 and ICP27. Interestingly, NEAT1_2 circular and linear splicing was also up-regulated in influenza A virus (IAV) infection but absent in stress conditions, which disrupt transcription termination similar to but not by the same mechanisms as HSV-1 and IAV infection. Large-scale analysis of published RNA-seq data uncovered induction of NEAT1_2 splicing in cancer cells upon inhibition or knockdown of cyclin-dependent kinase 7 (CDK7) or the MED1 subunit of the Mediator complex phosphorylated by CDK7. Finally, CDK7 inhibition also disrupted transcription termination, highlighting a possible link between disruption of transcription termination and NEAT1_2 splicing.

bioinformatics↗

bioRxiv2021

Herpes simplex virus 1 inhibits phosphorylation of RNA polymerase II CTD serine-7

Transcriptional activity of RNA polymerase II (Pol II) is orchestrated by post-translational modifications of the C-terminal domain (CTD) of the largest Pol II subunit, RPB1. Herpes Simplex Virus type 1 (HSV-1) usurps the cellular transcriptional machinery during lytic infection to efficiently express viral mRNA and shut down host gene expression. The viral immediate-early protein ICP22 interferes with serine 2 phosphorylation (pS2) of the Pol II CTD by targeting CDK9. The functional implications of this are poorly understood. Here, we report that HSV-1 also induces a global loss of serine 7 phosphorylation (pS7). This effect was dependent on the expression of the two viral immediate-early proteins, ICP22 and ICP27. While lytic HSV-1 infection results in efficient Pol II degradation late in infection, we show that pS2/S7 loss precedes the drop in Pol II level. Interestingly, mutation of the RPB1 polyubiquitination site mutation K1268, which prevents proteasomal RPB1 degradation during transcription-coupled DNA repair, displayed loss of pS2/S7 but retained much higher overall RPB1 protein levels even at late times of infection, indicating that this pathway mediates bulk Pol II protein loss late in infection but is not involved in early CTD dysregulation. Using -amanitin-resistant CTD mutants, we observed differential requirements for Ser2 and Ser7 for production of viral proteins, with Ser2 facilitating viral immediate-early gene expression and Ser7 appearing dispensable. Despite dysregulation of CTD phosphorylation and different requirements for Ser2/7, all CTD modifications tested could be visualized in viral replication compartments by immunofluorescence. These data expand the known means that HSV-1 employs to create pro-viral transcriptional environments at the expense of host responses.

molecular biology↗

bioRxiv2019

Integrative functional genomics decodes herpes simplex virus 1

Since the genome of herpes simplex virus 1 (HSV-1) was first sequenced more than 30 years ago, its predicted 80 genes have been intensively studied. Here, we unravel the complete viral transcriptome and translatome during lytic infection with base-pair resolution by computational integration of multi-omics data. We identified a total of 201 viral transcripts and 284 open reading frames (ORFs) including all known and 46 novel large ORFs. Multiple transcript isoforms expressed from individual gene loci explain translation of the vast majority of novel viral ORFs as well as N-terminal extensions (NTEs) and truncations thereof. We show that key viral regulators and structural proteins possess NTEs, which initiate from non-canonical start codons and govern subcellular protein localization and packaging. We validated a novel non-canonical large spliced ORF in the ICP0 locus and identified a 93 aa ORF overlapping ICP34.5 that is thus also deleted in the FDA-approved oncolytic virus Imlygic. Finally, we extend the current nomenclature to include all novel viral gene products. Taken together, this work provides a valuable resource for future functional studies, vaccine design and oncolytic therapies.

microbiology↗

bioRxiv2019

scSLAM-seq reveals core features of transcription dynamics in single cells

Current single-cell RNA sequencing approaches gives a snapshot of a cellular phenotype but convey no information on the temporal dynamics of transcription. Moreover, the stochastic nature of transcription at molecular level is not recovered. Here, we present single-cell SLAM-seq (scSLAM-seq), which integrates metabolic RNA labeling, biochemical nucleoside conversion and single-cell RNA-seq to directly measure total transcript levels and transcriptional activity by differentiating newly synthesized from pre-existing RNA for thousands of genes per single cell. scSLAM-seq recovers the earliest virus-induced changes in cytomegalovirus infection and reveals a so far hidden phase of viral gene expression comprising promiscuous transcription of all kinetic classes. It depicts the stochastic nature of transcription and demonstrates extensive gene-specific differences. These range from stable transcription rates to on-off dynamics which coincide with gene-/promoter-intrinsic features (Tbp-TATA-box interactions and DNA methylation). Gene but not cell-specific features thus explain the heterogeneity in transcriptomes between individual cells and the transcriptional response to perturbations.

genomics↗

bioRxiv2018

P-TEFb activation by RBM7 shapes a pro-survival transcriptional response to genotoxic stress

Cellular DNA damage response (DDR) involves dramatic transcriptional alterations, the mechanisms of which remain ill-defined. Given the centrality of RNA polymerase II (Pol II) promoter-proximal pause release in transcriptional control, we evaluated its importance in DDR. Here we show that following genotoxic stress, the RNA-binding motif protein 7 (RBM7) stimulates Pol II elongation and promotes cell viability by activating the positive transcription elongation factor b (P-TEFb). This is mediated by genotoxic stress-enhanced binding of RBM7 to 7SK snRNA (7SK), the scaffold of the 7SK small nuclear ribonucleoprotein (7SK snRNP) which inhibits P-TEFb. In turn, P-TEFb relocates from 7SK snRNP to chromatin to induce transcription of short units including key DDR genes and multiple classes of non-coding RNAs. Critically, interfering with RBM7 or P-TEFb provokes cellular hypersensitivity to DNA damage-inducing agents through activation of apoptotic program. By alleviating the inhibition of P-TEFb, RBM7 thus facilitates Pol II elongation to enable a pro-survival transcriptional response that is crucial for cell fate upon genotoxic insult. Our work uncovers a new paradigm in stress-dependent control of Pol II pause release, and offers the promise for designing novel anti-cancer interventions using RBM7 and P-TEFb antagonists in combination with DNA-damaging chemotherapeutics.

biochemistry↗