Biology subjects

Grundhoff, A.

Publications and source records attributed to Grundhoff, A..

At least 19 recordsLinked to original sources

bioRxiv2026

Phenotypic Screening Identifies Small-Molecule Inhibitors with Distinct Activities across the BK Polyomavirus Life Cycle

BACKGROUND: BK polyomavirus (BKPyV) reactivation is a major complication in kidney and hematopoietic stem cell transplant recipients, yet no specific antiviral therapy is currently available. Antiviral discovery is complicated by the restricted tropism and slow replication kinetics of BKPyV and its extensive dependence on cellular processes. RESULTS: We established a phenotypic high-throughput screening and validation pipeline to identify small molecule inhibitors of BKPyV infection. Using an SV40-infected CV1 reporter system, approximately 28,000 small molecules were screened, yielding 98 primary candidates. Confirmatory testing identified 33 compounds with reproducible activity, of which 16 subsequently inhibited BKPyV in human renal proximal tubular epithelial cells. Concentration response and cytotoxicity analyses revealed distinct antiviral potency and selectivity profiles, and integration of these data with predicted toxicity, physicochemical properties, and synthetic accessibility enabled further compound prioritization. Time of addition experiments revealed distinct temporal windows of antiviral activity, and MOI dependent concentration response analyses demonstrated that the potency of selected inhibitors varied with viral inoculum. Further characterization of prioritized compounds identified differential effects on BKPyV attachment and viral gene expression. Transcriptomic profiling of three selected compounds C5, C8, and C9 revealed distinct compound-associated cellular responses, supporting interference with different host-dependent processes during BKPyV infection. CONCLUSIONS: We identified a pharmacologically diverse panel of small-molecule inhibitors active against BKPyV in human renal epithelial cells. Their distinct potency, selectivity, temporal activity, and cellular response profiles indicate multiple modes of antiviral interference and establish C5, C8, and C9 as candidates for further target identification and optimization. More broadly, our findings demonstrate the utility of surrogate phenotypic screening for discovering inhibitors of BKPyV and provide new chemical tools to investigate host dependencies of the BKPyV life cycle.

microbiology↗

bioRxiv2026

Temporal Profiling of Upper-Layer Neurons Reveals Changes in the Molecular Landscape Upon Maternal Immune Activation

Neuronal differentiation is a dynamic, multi-layered process that transforms progenitor cells into functionally integrated neurons, yet the coordinated molecular programs underlying this transition remain incompletely understood. Here, we define the developmental trajectory of murine upper-layer cortical neurons using an integrated multi-omics approach combining transcriptomics and proteomics across key stages of neurogenesis. We find that neuronal identity emerges gradually through coordinated transitions from RNA processing and splicing programs toward synaptic and metabolic maturation. Integration of matched transcriptomic and proteomic datasets reveals a compact set of concordant molecular modules that define this trajectory, highlighting post-transcriptional regulation as a central driver of neuronal maturation. We further show that maternal immune activation (MIA), a model of prenatal inflammation, deranges this developmental program. MIA induces sustained upregulation of Wnt signalling pathways alongside a downregulation of synaptic regulators, without detectable global alterations in DNA methylation. These molecular changes are accompanied by defects in neuronal positioning during cortical development, linking altered molecular trajectories to functional outcomes. Together, our findings establish a temporal molecular framework of neuronal differentiation and demonstrate that prenatal environmental perturbations reshape cortical development primarily through post-transcriptional and signalling-based mechanisms.

neuroscience↗

bioRxiv2025

Defective proviruses cause T cell reprogramming through promoter exaptation in HIV-1 infection

People living with HIV (PLWH) on antiretroviral therapy (ART) accumulate primarily defective proviral sequences in genomes of often clonally expanded CD4+ HIV-1 target cells. The majority of viral-derived DNA is transcriptionally active and preferentially found at distinct genomic loci suggesting a selective process driven by integration site-specific crosstalk between viral and host sequences. Focusing on one of the most prominent selected integration loci, the BTB Domain and CNC Homolog 2 (BACH2) gene, we here show mechanistic insights how CD4+ T cells are functionally reprogrammed via exaptation of provirus-derived regulatory sequences during long-term ART. Using a cellular model of BACH2-integrated proviruses, we find that proviral transcription drives aberrant BACH2 protein levels that escape autoregulatory feedback and impose BACH2-dependent transcriptomic changes. By mimicking these changes in primary CD4+ T lymphocytes, we observe that BACH2 drives reprogramming of cells toward a proliferative, precursor memory-like type. These reprogrammed CD4+ T cells possess traits of immune evasion and cellular survival that are signatures of persistent HIV reservoir cells in PLWH. Inhibition of provirus transcriptional activity can mitigate exaptation, suggesting a strategy to offset HIV-driven differentiation and expansion of CD4+ T cells. Finally, our data suggest that provirus exaptation at a second prominently selected proviral integration gene, the Signal Transducer And Activator of Transcription 5B (STAT5B) gene, drives a contrary, effector-like T cell fate, suggesting a multifaceted impact of exaptation on immune homeostasis. Overall, our data suggest that transcriptionally active proviruses, even if structurally defective, modulate target cells through insertional activation of integration genes, a process which we postulate to contribute to the complex immune modulation and dysregulation experienced by ART-suppressed PLWH.

microbiology↗

bioRxiv2025

Oncogenic virus hijacks SOX18 pioneer function to enhance viral persistence

Kaposis sarcoma herpesvirus (KSHV) establishes lifelong oncogenic infection in lymphatic endothelial cells (LECs) by ensuring episomal maintenance of its genome via the viral protein LANA. Efficient viral genome maintenance typically involves host DNA replication and episome tethering, but the extent of cell-type-specific regulation remains unclear. Here, we identify that KSHV hijacks the pioneering function of the endothelial-specific transcription factor SOX18 to facilitate persistence of viral episomes. Upon infection, LANA co-opts SOX18 to recruit the SWI/SNF chromatin-remodeling complex via its ATPase subunit BRG1, enhancing chromatin accessibility and enabling efficient viral genome persistence. Disruption of SOX18 or BRG1--genetically or pharmacologically--leads to reduced episome load and attenuated hallmarks of virus infection. This work highlights how viruses can harness lineage-specific transcriptional regulators to establish persistent nuclear retention of their episome into the host genome.

molecular biology↗

bioRxiv2025

Merkel cell polyomavirus infection and persistence modelled in skin organoids

Merkel cell polyomavirus (MCPyV) causes most Merkel cell carcinomas (MCC). The virus is one of the few known human tumor viruses, and due to its direct role in this skin cancer development, it is a bona fide model for viral tumorigenesis and tumorigenesis in general. Chronic viruses in humans such as MCPyV are highly adapted to their host and current models to study infection, persistence and pathogenesis are highly limited. We here use an induced pluripotent stem cell (iPSC)-derived hair-bearing skin organoid (SkO) system to demonstrate efficient viral infection, progression and spread of MCPyV. Using bulk-, single cell - and spatial-transcriptomics, combined with immunostaining and nucleic acid hybridization technologies, we show that MCPyV ensures persistence due to a quasi-latency state within the majority of dermal fibroblasts carry the viral genome. Further, we identify the cell type of productive infection with papillary fibroblasts and dermal sheath fibroblasts supporting viral replication and progeny production. Our high-resolution methods demonstrate that the virus in these cells evades the innate immune response, as evidenced by the efficacy of interferon-beta treatment or ruxolitinib, a JAK/STAT inhibitor, in suppressing or stimulating viral replication. We show that iPSC-derived SkOs are able to support infection and long-term persistence of the virus under conditions very similar to those found in humans. Thus, this infection model provides a robust platform for understanding and characterizing the interaction of this virus with the immune system in infection, testing treatment strategies to control reactivation and map processes involved in tumor development.

molecular biology↗

bioRxiv2025

Complex Human Hair Bearing Skin Organoids as Model for Herpes Simplex Virus 1 Infection in the Skin

Epithelial surfaces are the initial site of Herpes Simplex Virus 1 (HSV-1) infection. After latency establishment in the peripheral nervous system, reactivation can lead to skin pathologies. To reflect the complexity of skin we used complex human hair bearing skin organoids as a new model system for HSV-1 infection. We analyzed spread of the infection through the three-dimensional tissue and the cell type specific host response in a spatial temporal manner. Single-cell-resolution spatial transcriptomics and imaging shows a restricted viral infection in keratinocytes of the epidermis and hair follicles. The host factor IER3 is upregulated in papillary fibroblasts of the dermis as the infection progresses through the tissue. In keratinocytes of the stratum basale, we find IRF3 upregulated in infected but also virus negative cells, suggesting a signaling mechanism to initiate antiviral responses. Furthermore, we find a cell type specific inflammatory response with an upregulation of TNFSF9 in dermal fibroblasts and TNF in keratinocytes. TNF target genes are induced in neighboring uninfected cells suggesting the induction of a localized inflammatory reaction to suppress viral infection. In summary, our findings position human hair-bearing skin organoids as a highly physiological model system for studying HSV-1 pathogenesis and reveal spatially organized host responses.

microbiology↗

bioRxiv2025

Polycomb group proteins protect latent Kaposi sarcoma-associated herpesvirus from episome clearance and HUSH-dependent chromatin silencing

Kaposi sarcoma-associated herpesvirus (KSHV) is the causative agent of a number of human neoplasms. KSHV causes lifelong chronic infections by establishing persistent reservoirs of latently infected cells in which the viral double-stranded DNA genome is present in the form of an extrachromosomal episome. Polycomb group protein (PcG) mediated gene repression is detected early after entry of the CpG-rich and epigenetically naive viral DNA into the host nucleus. Here we show that the establishment and maintenance of KSHV latency is strongly influenced by Polycomb Repressive Complexes 1 and 2 (PRC1, and PRC2, respectively). Using a series of genetic deletion models, we dissect the functional interdependence of PRC1 and PRC2 on KSHV and identify KSHV episomes as a prime target for independent as well as collaborative de novo PRC recruitment. Loss of PRC function leads to a severely altered chromatin state on KSHV and we show that PRC mediated gene repression is not only preventing transcription of viral genomes but also protects KSHV episomes from detection by the human silencing hub (HUSH). Furthermore, we demonstrate that PRC complexes play an important role in ensuring episomal maintenance in dividing cells. Taken together, our results show that absence of PRC complexes creates a metastable state of KSHV latency and suggest early chromatinization as an integral step for KSHV latency establishment.

microbiology↗

bioRxiv2025

Spatiotemporal analysis of de novo KSHV infection using Crispr/Cas9-based 3D live cell imaging at single episome resolution

Kaposi Sarcoma-associated herpesvirus (KSHV) persists as a latent episome in infected cells. While the virus efficiently infects established cell lines and primary cells in vitro, the early events guiding establishment of latent infection and the dynamic interplay between viral episomes and host factors remain incompletely understood. Here, we describe the development and application of a CRISPR/Cas9-based 3D live cell imaging system capable of tracking single KSHV episomes in real-time. Our approach exploits the SunTag technology, wherein deactivated Cas9 (dCas9) molecules are fused to repetitive epitope arrays recognized by superfolder GFP-fused single-chain antibodies. By targeting these complexes to terminal repeat units of KSHV, we achieve high level signal amplification, allowing us not only to detect newly incoming viral genomes within the first hours of de novo infection, but also to follow their spatiotemporal trajectories through different stages of the viral lifecycle. Furthermore, to facilitate efficient generation of stable reporter cell lines, we adapted the transposon-based piggyBac system to combine all SunTag components into a single-vector targeting system (SunSeT). Using these systems, we demonstrate the ability to observe both transient and stable interactions between KSHV episomes and key cellular regulators, including the variant polycomb-repressive complex 1 (vPRC) component KDM2B and the innate immune sensor IFI16. Furthermore, our platform allows detailed visualization of episodic changes in episome localization, abundance and distribution during de novo and long-term infection, providing critical insights into how viral genome positioning and dynamics correlate with host subnuclear environments. Overall, our study introduces a robust and adaptable imaging platform to dissect the earliest events of KSHV infection. The ability to track viral episomes in living cells offers a powerful tool to advance our understanding of the spatial and temporal regulation of individual KSHV genomes, shedding light on fundamental mechanisms of herpesvirus latency and persistence.

microbiology↗

bioRxiv2025

Intrinsic immunity against HAdV is achieved by a novel epigenetic silencing complex

The DNA double-stranded genome of human adenoviruses (HAdV) is preferentially targeted by host factors involved in chromatin remodeling, such as SPOC1 and KAP1 to inhibit efficient viral gene expression. HAdV genomes undergo alterations through association with host histones and epigenetic modifications; however, the precise underlying mechanism remains elusive. A recently discovered silencing mechanism for retrotransposons and retroviruses involves the Human Silencing Hub (HUSH) complex. This complex of MPP8, TASOR, and PPHLN1 safeguards the human genome by utilizing histone H3 Lys9 trimethylation (H3K9me3) to block transcription. Through the recruitment of SETDB1 and MORC2, the HUSH complex silences host genes and condenses target genomes to combat infections such as HIV, MLV, and AAV. Here, we present evidence that the HUSH complex effectively restricts HAdV infection. To counteract the repressive function of this epigenetic silencing complex, HUSH factors are inhibited through binding of HAdV proteins and subsequent relocalization. We observe that MPP8 is targeted by the adenoviral E3 ubiquitin ligase, thus recruited by the viral early proteins E1B-55K and E4orf6 for proteasomal degradation. In summary, we provide evidence that the HUSH complex is a previously unrecognized host factor that restricts HAdV gene expression and replication. Based on these novel findings, we propose that HUSH represents a promising therapeutic target to combat HAdV infection.

cell biology↗

bioRxiv2025

E2A SUMOylation promotes HAdV mediated inhibition of the tumor suppressor p53 during infection

HAdV express early viral genes to modulate the activity of the cellular tumor suppressor p53 and ensure efficient viral replication, through various processes, whilst some of them are not completely understood. HAdV oncoprotein E1B-55K interactions with SUMO, PML-IV/V and Sp100A are tightly connected to the transforming potential of the viral factor in non-lytic infections. SUMO modified HAdV DNA binding protein (DBP) E2A interacts with PML and Sp100A acting as a molecular bridge between viral replication centers (RCs) and PML tracks. Here, we provide a novel concept showing that HAdV E2A is involved in the inhibition of p53 activity and apoptosis; thus we identified a novel route of exhibiting virus induced oncogenic potential. We revealed an E1B-55K independent localization of p53 within HAdV replication centers marked by E2A. In the absence of any functional E1B-55K protein expression, p53 interacts with E2A. E2A SUMOylation promotes HAdV- mediated inhibition of p53/DNA binding and p53-dependent transactivation by supporting p53 degradation, E1B-55K interaction and colocalization with PML-V that leads to E1B-55K induced upregulation of p53 SUMOylation. In sum, we provide evidence for a novel mechanism by which the HAdV DBP E2A protein inhibits p53- mediated transcriptional activation and apoptotic host cell response. Intriguingly, these findings change the longstanding dogma of host DNA damage response (DDR) inactivation by HAdV infection and offer crucial insights to improve future p53 selective oncolytic virus therapies and adenoviral vectors.

cell biology↗

bioRxiv2024

Repression of CADM1 transcription by HPV type 18 is mediated by three-dimensional rearrangement of promoter-enhancer interactions

Upon infection, human papillomavirus (HPV) manipulates host cell gene expression to create an environment that is supportive of a productive and persistent infection. The virus-induced changes to the host cells transcriptome are thought to contribute to carcinogenesis. Here, we show by RNA-sequencing that oncogenic HPV18 episome replication in primary human foreskin keratinocytes (HFKs) drives host transcriptional changes that are consistent between multiple HFK donors. We have previously shown that HPV18 episome replication in HFKs results in post-transcriptional stabilisation of the host chromatin insulation protein CTCF. Since CTCF is an important regulator of host cell transcription via coordination of epigenetic boundaries and long-range chromosomal interactions, we hypothesised that HPV18-induced stabilisation of CTCF may contribute to host transcription reprogramming. Analysis of CTCF binding in the host cell genome by ChIP-Seq revealed that while the total number of CTCF binding sites is not altered by the virus, there are a sub-set of CTCF binding sites that are either enriched or depleted of CTCF. Many of these altered sites are clustered within regulatory elements of differentially expressed genes, including the tumour suppressor gene cell adhesion molecule 1 (CADM1), which supresses epithelial cell growth and invasion. We show that HPV18 establishment results in reduced CTCF binding at the CADM1 promoter and upstream enhancer. Loss of CTCF binding is coincident with epigenetic repression of CADM1, in the absence of CpG hypermethylation, while adjacent genes including the transcriptional regulator ZBTB16 are activated. These data indicate that the CADM1 locus is subject to topological rearrangement following HPV18 establishment. We tested this hypothesis using 4C-Seq (circular chromosome confirmation capture-sequencing) and show that HPV18 establishment causes a loss of long-range chromosomal interactions between the CADM1 transcriptional start site and the upstream transcriptional enhancer. These data show that HPV18 manipulates host cell promoter-enhancer interactions to drive transcriptional reprogramming that may contribute to HPV-induced disease progression. AUTHOR SUMMARYInfection with oncogenic HPV is the cause of numerous cancer types, which generally arise after persistent HPV infection. Upon infection, HPV alters the gene expression profile of infected cells to facilitate virus replication and persistence. Multiple mechanisms of HPV-induced host cell reprogramming have been previously suggested. Here, we show that HPV infection induces rearrangement of specific genomic loci by altering the chromatin binding of the host cell protein CTCF, an important regulator of chromatin architecture. Loss of CTCF binding to a cluster of binding sites at the CADM1 locus on chromosome 11 is coincident with epigenetic reprogramming and disruption of long-range chromatin interactions, resulting in transcriptional repression of CADM1. Our data show that repression of CADM1 is an early event in HPV-driven disease, preceding hypermethylation of the CADM1 transcriptional promoter that is frequently observed in HPV-driven cancers, demonstrating a novel mechanism of HPV-induced host cell transcriptional reprogramming.

microbiology↗

bioRxiv2024

Limited protection against early-life cytomegalovirus infection results from deficiency of cytotoxic CD8T cells

Differential antiviral T cell immunity in early life impacts the clinical outcome of Cytomegalovirus (CMV) infection, but the underlying mechanisms are not well understood. Here, we found delayed enrichment of early-life murine CMV-specific CD8 T cells due to a general deficiency of {beta} T cells. Adoptive transfer of naive adult T cells into neonates did not protect due to a blockade of CD8 but not of CD4 effector T cell differentiation. Early-life deficiency of critical signal 3 cytokines during T cell priming resulted in the appearance of non-cytotoxic CD8 effector T cells whereas the effector phase of adult-primed T cells was not disrupted in neonates. Accordingly, we found an overall low number of antiviral human CD8 T cells in newborns with congenital CMV. Together, this study suggests defective CD8 T cell immunity as an important factor explaining the higher risk for CMV disease in the early-life phase.

immunology↗

bioRxiv2024

Merkel cell polyomavirus small tumor antigen contributes to immune evasion by interfering with type I interferon signaling

Merkel cell polyomavirus (MCPyV) is the causative agent of the majority of Merkel cell carcinomas (MCC). The virus has limited coding capacity, with its early viral proteins, large T (LT) and small T (sT), being multifunctional and contributing to infection and transformation. A fundamental difference in early viral gene expression between infection and MCPyV-driven tumorigenesis is the expression of a truncated LT (LTtr) in the tumor. In contrast, sT is expressed in both conditions and contributes significantly to oncogenesis. Here, we identified novel functions of early viral proteins by performing genome-wide transcriptome and chromatin studies in primary human fibroblasts. Due to current limitations in infection and tumorigenesis models, we mimic these conditions by ectopically expressing sT, LT or LTtr, individually or in combination, at different time points. In addition to its known function in cell cycle and inflammation modulation, we reveal a fundamentally new function of sT. We show that sT regulates the type I interferon (IFN) response downstream of the type I interferon receptor (IFNAR) by interfering with the interferon-stimulated gene factor 3 (ISGF3)-induced interferon-stimulated gene (ISG) response. Expression of sT leads to a reduction in the expression of interferon regulatory factor 9 (IRF9) which is a central component of the ISGF3 complex. We further show that this function of sT is conserved in BKPyV. We provide a first mechanistic understanding of which early viral proteins trigger and control the type I IFN response, which may influence MCPyV infection, persistence and, during MCC progression, regulation of the tumor microenvironment. Author SummaryMerkel cell polyomavirus (MCPyV) is the only human polyomavirus that causes cancer in humans. As with all human polyomaviruses, the available infection models are limited. Thus, many processes such as the host response to infection and its regulation by the virus to establish infection and persistence are poorly understood. To better understand this interplay of viral MCPyV proteins, we performed genome-wide transcriptome and chromatin studies in primary human fibroblasts and simulated infection and tumorigenesis conditions by ectopically expressing the early viral proteins individually or in combination at different time points. This allowed us to uncover a novel, previously undescribed function of polyomavirus sT, namely the reduction of the ISG response by affecting the ISGF3 complex, specifically by reducing IRF9 protein levels. This work sheds light on how early viral proteins influence the type I IFN response and how their interplay may affect MCPyV infection, persistence, and MCC progression.

molecular biology↗

bioRxiv2024

Repression of varicella zoster virus gene expression during quiescent infection in the absence of detectable histone deposition

Varicella zoster virus (VZV) is a human-specific herpesvirus that establishes latency in peripheral neurons. The only transcripts detected in infected human trigeminal ganglia (TG) obtained shortly after death correspond to the VZV latency-associated transcript (VLT) and associated VLT-ORF63 splice variants. In vitro studies showed that VLT-ORF63 is translated into a protein (pVLT-ORF63) that induces VZV transcription. The mechanisms that lead to this restricted gene expression and the transition to lytic replication remain unknown, partly due to the difficulty of working with human neurons. In this study, we addressed whether the neuroblastoma-derived cell line SH-SY5Y could serve as a model to investigate the mechanisms that lead to repression of VZV gene expression followed by reactivation. VZV productively infected differentiated SH-SY5Y (dSH-SY5Y) whereas incubation with acyclovir (ACV) inhibited virus replication and induced a progressive repression of the virus. Upon removal of ACV there was production of viral particles in a subset of cells, while others contained non-replicating VZV genomes and VLT-containing transcripts for at least 20 days post-infection (dpi). Exogenous expression of VLT-ORF63 induced productive infection, suggesting that the non-replicating and repressed genomes remained functional. Interestingly, histone deposition was undetectable at VZV genomes in quiescently infected dSH-SY5Y cells, pointing to a potential novel mechanism leading to VZV repression in this neuronal setting.

microbiology↗

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↗

bioRxiv2021

Epigenetic reprogramming of human macrophages by an intestinal pathogen

Various pathogens systematically reprogram gene expression in macrophages, but the underlying mechanisms are largely unknown. We investigated whether the enteropathogen Yersinia enterocolitica alters chromatin states to reprogram gene expression in primary human macrophages. Genome-wide chromatin immunoprecipitation (ChIP) seq analyses showed that pathogen-associated molecular patterns (PAMPs) induced up- or down-regulation of histone modifications (HMod) at approximately 14500 loci in promoters and enhancers. Effectors of Y. enterocolitica reorganized about half of these dynamic HMod, with the effector YopP being responsible for about half of these modulatory activities. The reorganized HMod were associated with genes involved in immune response and metabolism. Remarkably, the altered HMod also associated with 61 % of all 534 known Rho GTPase pathway genes, revealing a new level in Rho GTPase regulation and a new aspect of bacterial pathogenicity. Changes in HMod were associated to varying degrees with corresponding gene expression, e. g. depending on chromatin localization and cooperation of the HMod. In summary, infection with Y. enterocolitica remodels HMod in human macrophages to modulate key gene expression programs of the innate immune response. Author SummaryHuman pathogenic bacteria can affect epigenetic histone modifications to modulate gene expression in host cells. However, a systems biology analysis of this bacterial virulence mechanism in immune cells has not been performed. Here we analyzed genome-wide epigenetic histone modifications and associated gene expression changes in primary human macrophages infected with enteropathogenic Yersinia enterocolitica. We demonstrate that Yersinia virulence factors extensively modulate histone modifications and associated gene expression triggered by the pathogen-associated molecular patterns (PAMPs) of the bacteria. The epigenetically modulated genes are involved in several key pathways of the macrophage immune response, including the Rho GTPase pathway, revealing a novel level of Rho GTPase regulation by a bacterial pathogen. Overall, our findings provide an in-depth view of epigenetic and gene expression changes during host-pathogen interaction and might have further implications for understanding of the innate immune memory in macrophages.

microbiology↗

bioRxiv2020

Human antigen-specific memory natural killer cell responses develop against HIV-1 and influenza virus and are dependent on MHC-E restriction

For over a decade, multiple studies have disputed the notion of natural killer (NK) cells as purely innate lymphocytes by demonstrating that they are capable of putative antigen-specific immunological memory against multiple infectious agents including two critical global health priorities - HIV and influenza. However, the mechanisms underlying antigen specificity remain unknown. Herein, we demonstrate that antigen-specific human NK cell memory develops upon exposure to both HIV and influenza, unified by a conserved and epitope-specific targetable mechanism firmly dependent on the activating CD94/NKG2C receptor and its ligand HLA-E, and confirm these findings by three rigorous and novel assays. We validated the permanent acquisition of antigen-specificity by individual memory NK cells by single-cell cloning. We identified biomarkers of antigen-specific NK cell memory through RNA-Seq transcriptomic fingerprints and complex immunophenotyping by 30-parameter flow cytometry showing elevated expression of KLRG1, 4{beta}7 and NKG2C. Finally, we show individual HLA-E-restricted peptides that may constitute the dominant response in HIV-1- and influenza-infected persons in vivo. Our findings clarify the mechanisms behind formation of antigen-specific memory NK cells, and suggest they could be targeted for future vaccines, cure strategies, or other therapeutic interventions.

immunology↗

bioRxiv2020

SARS-CoV-2 infects carotid arteries: implications for vascular disease and organ injury in COVID-19

Stroke and central nervous system dysfunction are cardinal symptoms in critically ill corona virus disease 19 (COVID-19) patients. In an autopsy series of 32 COVID-19 patients, we investigated whether carotid arteries were infected with SARS-CoV-2 by employing genomic, virologic, histochemical and transcriptomic analyses. We show that SARS-CoV-2 productively infects and modulates vascular responses in carotid arteries. This finding has far reaching implications for the understanding and clinical treatment of COVID-19.

microbiology↗