Biology subjects

Witt, A.

Publications and source records attributed to Witt, A..

11 recordsLinked to original sources

bioRxiv2026

Cross-Continental Analysis of Vampire Bat Betaherpesvirus Reveals Limited Interference Among Strains and Local Geographic Spread

Desmodus rotundus Betaherpesvirus (DrBHV) is a candidate vector for a transmissible vaccine targeting the circulation of rabies virus within its vampire bat reservoir. Studies assessing the potential for DrBHV as a vector have not considered its geographic range, potential for transboundary spread or how the diversity of wildtype DrBHV in natural bat populations might impede the spread of a modified vaccine strain. Here, by sequencing DrBHVs from vampire bats spanning 15 regions across 7 rabies-affected countries in Latin America and the Caribbean, we characterise the continent-scale distribution of DrBHV diversity and demonstrate widespread and apparently unconstrained co-infection. DrBHV occurred in all regions, forming a monophyletic clade consistent with a single introduction to vampire bats or host-virus co-speciation rather than frequent host switching. Phylogeographic analyses revealed cross-boundary spread that was predicted by geographic proximity. We identified 50 putative DrBHV strains and 79% of DrBHV-infected bats harboured multiple strains. No strains were over- or under-represented in co-infection and co-infections occurred proportionately to local strain prevalence. Whether strains circulated in given populations was predominantly driven by the geographic proximity of other populations containing that strain. The evolutionary relatedness of strains constrained neither rates of co-infection within individuals nor whether strains co-circulated within regions, suggesting vaccine vectors might be locally sourced rather than requiring imported, divergent strains to ameliorate interference. These results support the viability of DrBHV-vectored vaccines for mitigating rabies virus across Latin America and the Caribbean, despite widespread circulation of wildtype viruses.

genomics↗

bioRxiv2025

Differentiation-Dependent Proximity Proteomics Identifies Novel Host Factors Linked to HPV16 E2 Function

Human papillomavirus 16 (HPV16) is a causative agent of oropharyngeal, cervical and anogenital cancers. The viral E2 protein is essential for viral genome replication, transcriptional regulation, episome maintenance, and activation of the host DNA damage response. Despite its central role, the full network of HPV16 E2 interactions with host proteins remains incompletely defined, particularly under differentiating conditions which support the complete viral life cycle. In this study, we used TurboID-based proximity labeling to characterize the interactome of HPV16 E2 and known host partner protein TOPBP1, in both undifferentiated monolayer and differentiating keratinocytes. We generated stable keratinocyte lines expressing doxycycline-inducible TurboID-tagged HPV16 E2 and confirmed that the tagged protein retained transcriptional, replicative, and DNA damage-inducing functions. Mass spectrometry analysis of streptavidin-enriched proteins identified both known and novel E2-associated host factors, including chromatin regulators, DNA repair proteins, and nucleolar components. Comparative analysis revealed a substantial overlap between E2 and TOPBP1 interactomes, and in situ validation by proximity ligation assay identified nucleolin (NCL) as a differentiation-dependent factor whose interaction with E2 is stabilized by TOPBP1. Functional studies demonstrated that NCL is required for episomal genome maintenance, highlighting a cooperative E2-TOPBP1- NCL axis critical for viral genome stability during differentiation. These findings provide a comprehensive view of the E2-associated protein landscape in stratified epithelial cells and reveal a mechanistic pathway through which HPV16 co-opts host factors to support genome maintenance, productive replication, and persistence. ImportanceHuman papillomaviruses (HPVs) establish persistent infections in stratified epithelia and rely on host DNA damage and repair factors to support their replication. The E2 protein is central to viral genome replication and maintenance, and depends heavily on its interaction with the host factor TOPBP1 for these functions. Here, we define the E2 and TOPBP1 interactomes in differentiating keratinocytes, and identify nucleolin (NCL) a critical differentiation- and TOPBP1-dependent E2 partner required for episomal genome stability. These findings expand the understanding of how HPV16 coordinates viral replication with host chromatin and DNA repair networks, uncovering a cooperative E2-TOPBP1-NCL axis that may represent a new target for antiviral intervention.

cancer biology↗

bioRxiv2025

E2 displacement of CIP2A from TOPBP1 activates the DNA damage response during papillomavirus life cycles

The papillomavirus life cycle is intricately linked to epithelial differentiation, and the virus manipulates the differentiation process to facilitate viral production. One such manipulation is activation of the DNA damage response (DDR) which promotes viral replication via homologous recombination. This report demonstrates that the papillomavirus transcription/replication/segregation factor E2 activates the DNA damage response (DDR). During differentiation, E2 displacement of CIP2A from TOPBP1 causes CIP2A to bind and inhibit PP2A resulting in DDR activation via ATM phosphorylation. The DDR promotes inhibitory interaction of DBC1 with the class III deacetylase SIRT1, which further boosts the DDR via increased acetylation and stability of viral and host proteins. E2 forms a complex with TOPBP1 and ATM, while preventing ATR activation by blocking TOPBP1-ATR interaction. This "ATM up ATR down" phenotype promotes viral replication via ATM promotion of homologous recombination, and cell proliferation via inhibition of ATR. We demonstrate this mechanism of DDR activation in multiple systems: keratinocytes expressing only E2, in foreskin keratinocytes immortalized by HPV16, in HPV16 positive keratinocytes derived from a cervical lesion, in pre-neoplastic lesions induced by mouse papillomavirus MmuPV1, in head and neck cancer cell lines that retain E2 expression, and in HPV16 positive oropharyngeal patient derived xenografts that retain E2 expression. ATM inhibition preferentially killed cells expressing E2, presenting a novel strategy for treating HPV early preneoplasia and a large subset of HPV+ oropharyngeal cancers retaining E2 expression and episomal genomes.

cancer biology↗

bioRxiv2025

HPV16 recruitment of SMARCAL1 to viral and host replication forks is required for the viral life cycle

High-risk human papillomaviruses (HR HPVs) are responsible for around 5% of the worlds cancer burden. Activation and interaction with the host DNA damage response (DDR) promotes the HPV16 life cycle. This study demonstrates a crucial interaction between HPV16 and SMARCAL1, a protein involved in the stabilization of stalled DNA replication forks. SMARCAL1 can complex with E2, is recruited to E1-E2 replicating DNA, and SMARCAL1 knockdown reduces the fidelity of E1-E2 mediated DNA replication in C33a cells but does not alter replication levels. SMARCAL1 is recruited to the HPV16 genome in HPV16-immortalized foreskin keratinocytes (HFK+HPV16), and in situ protein interaction with nascent DNA replication forks (SIRF) assays demonstrated that SMARCAL1 is hyper-recruited to host replication forks in HFK+HPV16 cells. Using COMET, DNA fiber, and cell growth assays it was determined that knockdown of SMARCAL1 increased DNA damage and impaired replication fork progression in HFK+HPV16 cells, ultimately resulting in growth arrest. The viral genome integrates following SMARCAL1 knockdown in HFK+HPV16 cells. Therefore, SMARCAL1 facilitates host and viral DNA replication in HFK+HPV16 cells. Overall, the results demonstrate that HPV16 promotes SMARCAL1 recruitment to viral and host replication forks and is an essential factor for the HPV16 life cycle. The results expand our understanding of DDR proteins that regulate the HPV16 life cycle, and suggest that inhibition of SMARCAL1 function represents a novel anti-viral strategy for the treatment and prevention of HPV infections. ImportanceHPV16 is responsible for the majority of HPV+ cancers, contributing to 54% of cervical cancers and [~]90% of HPV+HNSCC. Integration of viral genomes into host DNA can promote cervical cancer progression and correlates with poor prognosis in HPV-associated HNSCC, where around 70% of HPV+ cancers contain episomal viral genomes. Developing effective antiviral therapies requires a deeper understanding of the interplay between viral replication and host DNA damage response (DDR) pathways. This report demonstrates that SMARCAL1 is essential for HPV16 replication and keratinocyte proliferation and that its depletion leads to replication stress, DNA damage, and viral genome integration. This work underscores the delicate balance between viral exploitation of the host DDR and the risk of genome instability. These insights contribute to the broader understanding of HPV pathogenesis and may inform the development of therapeutic strategies targeting viral replication to prevent disease progression and improve clinical outcomes in HPV-associated cancers.

microbiology↗

bioRxiv2025

Movement sonification during haptic exploration shifts emotional outcome without altering texture perception

Adding movement sonification to haptic exploration can change the perceptual outcome of a textured surface through multisensory processing. We hypothesized that auditory-evoked emotions would influence the appraisal of textured surfaces, with corresponding changes reflected in cortical excitability. Participants actively rubbed two different textured surfaces (slippery and rough) either without movement sonification, or with pleasant or disagreeable movement sonification. We found that sounds, whether agreeable or disagreeable, did not change the texture appraisal. However, the less pleasant surface was associated with a stronger negative hedonic valence, particularly when paired with disagreeable movement sonification. Time frequency analyses of EEG activities revealed a significant reduction in beta-band power [15-25 Hz] within the source-estimated sensorimotor and superior posterior parietal cortices when contrasting both pleasant and unpleasant sound with the silent touch. This suggests that the primary somatosensory cortices together with the superior parietal regions participated in the audio-tactile binding, with both pleasant and unpleasant sounds. In addition, we observed a significant increase in beta-band power in medial visual areas, specifically when disagreeable movement sonification was paired with tactile exploration. This may reflect a disengagement of visual cortical processing, potentially amplifying auditory-driven emotional responses and intensifying the perceived unpleasantness of the explored surfaces.

neuroscience↗

bioRxiv2024

Fibroblast Stromal Support Model for Predicting Human Papillomavirus-Associated Cancer Drug Responses.

Currently, there are no specific antiviral therapeutic approaches targeting Human papillomaviruses (HPVs), which cause around 5% of all human cancers. Specific antiviral reagents are particularly needed for HPV-related oropharyngeal cancers (HPV+OPCs) whose incidence is increasing and for which there are no early diagnostic tools available. We and others have demonstrated that the estrogen receptor alpha (ER) is overexpressed in HPV+OPCs, compared to HPV-negative cancers in this region, and that these elevated levels are associated with an improved disease outcome. Utilizing this HPV+ specific overexpression profile, we previously demonstrated that estrogen attenuates the growth and cell viability of HPV+ keratinocytes and HPV+ cancer cells in vitro. Expansion of this work in vivo failed to replicate this sensitization. The role of stromal support from the tumor microenvironment (TME) has previously been tied to both the HPV lifecycle and in vivo therapeutic responses. Our investigations revealed that in vitro co-culture with fibroblasts attenuated HPV+ specific estrogen growth responses. Continuing to monopolize on the HPV+ specific overexpression of ER, our co-culture models then assessed the suitability of the selective estrogen receptor modulators (SERMs), raloxifene and tamoxifen, and showed growth attenuation in a variety of our models to one or both of these drugs in vitro. Utilization of these SERMs in vivo closely resembled the sensitization predicted by our co-culture models. Therefore, the in vitro fibroblast co-culture model better predicts in vivo responses. We propose that utilization of our co-culture in vitro model can accelerate cancer therapeutic drug discovery. ImportanceHuman papillomavirus-related cancers (HPV+ cancers) remain a significant public health concern, and specific clinical approaches are desperately needed. In translating drug response data from in vitro to in vivo, the fibroblasts of the adjacent stromal support network play a key role. Our study presents the utilization of a fibroblast 2D co-culture system to better predict translational drug assessments for HPV+ cancers. We also suggest that this co-culture system should be considered for other translational approaches. Predicting even a portion of treatment paradigms that may fail in vivo with a co-culture model will yield significant time, effort, resource, and cost efficiencies.

cancer biology↗

bioRxiv2023

Human Papillomavirus 16 replication converts SAMHD1 into a homologous recombination factor and promotes its recruitment to replicating viral DNA

We have demonstrated that SAMHD1 (sterile alpha motif and histidine-aspartic domain HD-containing protein 1) is a restriction factor for the HPV16 life cycle. Here we demonstrate that in HPV negative cervical cancer C33a cells and human foreskin keratinocytes immortalized by HPV16 (HFK+HPV16), SAMHD1 is recruited to E1-E2 replicating DNA. Homologous recombination (HR) factors are required for HPV16 replication and viral replication promotes phosphorylation of SAMHD1, which converts it from a dNTPase to an HR factor independent from E6/E7 expression. A SAMHD1 phosphor-mimic (SAMHD1 T592D) reduces E1-E2 mediated DNA replication in C33a cells and has enhanced recruitment to the replicating DNA. In HFK+HPV16 cells SAMHD1 T592D is recruited to the viral DNA and attenuates cellular growth, but does not attenuate growth in isogenic HFK cells immortalized by E6/E7 alone. SAMHD1 T592D also attenuates the development of viral replication foci following keratinocyte differentiation. The results indicated that enhanced SAMHD1 phosphorylation could be therapeutically beneficial in cells with HPV16 replicating genomes. Protein phosphatase 2A (PP2A) can dephosphorylate SAMHD1 and PP2A function can be inhibited by endothall. We demonstrate that endothall reduces E1-E2 replication and promotes SAMHD1 recruitment to E1-E2 replicating DNA, mimicking the SAMHD1 T592D phenotypes. Finally, we demonstrate that in head and neck cancer cell lines with HPV16 episomal genomes endothall attenuates their growth and promotes recruitment of SAMHD1 to the viral genome. The results suggest that targeting cellular phosphatases has therapeutic potential for the treatment of HPV infections and cancers. ImportanceHuman papillomaviruses are causative agents in around 5% of all human cancers. The development of anti-viral therapeutics depends upon an increased understanding of the viral life cycle. Here we demonstrate that HPV16 replication converts SAMHD1 into an HR factor via phosphorylation. This phosphorylation promotes recruitment of SAMHD1 to viral DNA to assist with replication. A SAMHD1 mutant that mimics phosphorylation is hyper-recruited to viral DNA and attenuates viral replication. Expression of this mutant in HPV16 immortalized cells attenuates the growth of these cells, but not cells immortalized by the viral oncogenes E6/E7 alone. Finally, we demonstrate that the phosphatase inhibitor endothall promotes hyper-recruitment of endogenous SAMHD1 to HPV16 replicating DNA and can attenuate the growth of both HPV16 immortalized human foreskin keratinocytes and HPV16 positive head and neck cancer cell lines. We propose that phosphatase inhibitors represent a novel tool for combating HPV infections and disease.

cancer biology↗

bioRxiv2022

Inhibition of LATS Kinases in Ovarian Cancer Activates Cyclin D1/CDK4 and Decreases DYRK1A Activity

The controlled division of cells requires a coordination of multiple cellular pathways. Hippo pathway controls the organ size and restricts cell proliferation in response to the signals from cell surface receptors, and genetic alterations in the components of this pathway are common in cancer. LATS1 and LATS2 are homologous protein kinases that relay the signals from the environment to the Hippo effector YAP by direct phosphorylation that promotes its degradation. The genes encoding these kinases undergo frequent genetic losses in human cancers, with particularly high rates in high grade serous ovarian carcinoma (HGSOC), a highly lethal cancer with poorly understood mechanisms of pathogenesis and progression. We hypothesized that loss of LATS kinases could be a driver in this cancer and investigated signaling pathways downstream of LATS that could influence the ovarian cancer tumorigenic phenotypes. Depletion of both LATS1 and LATS2 was required to increase cell proliferation and disrupt the assembly of the cell-cycle regulatory DREAM complex. LATS-depleted human ovarian cancer cells formed bigger tumors in the immunocompromised mice, consistent with their tumor suppressor role. DREAM assembly depends on the activity of DYRK1A protein kinase, which was decreased in the LATS1/2-depleted cells. Furthermore, loss of LATS kinases increased the inhibitory phosphorylation of the retinoblastoma (Rb) family proteins, further promoting the DREAM disassembly that was rescued by CDK4 inhibitor palbociclib. Our study describes a crosstalk between the Hippo pathway and the cell cycle regulatory machinery converging on cyclin D1, a major regulator of the Rb tumor suppressor family, and highlights cellular pathways that could contribute to ovarian cancer pathogenesis and progression.

cancer biology↗

bioRxiv2022

INFLAMMATION INDUCED DOWNREGULATION OF SUCCINATE DEHYDROGENASE REGULATES ADRENOCORTICAL FUNCTION

The hypothalamus-pituitary-adrenal (HPA) axis is activated in response to inflammation leading to increased production of anti-inflammatory glucocorticoids by the adrenal cortex, thereby representing an endogenous feedback loop. However, severe inflammation reduces the responsiveness of the adrenal gland to adrenocorticotropic hormone (ACTH) although the underlying mechanisms are poorly understood. Here, we show by transcriptomic, proteomic and metabolomic analyses that LPS-induced systemic inflammation triggers profound metabolic changes in steroidogenic adrenocortical cells, including downregulation of the TCA cycle and oxidative phosphorylation. Inflammation disrupts the TCA cycle at the level of succinate dehydrogenase (SDH) leading to succinate accumulation and disturbed steroidogenesis. Mechanistically, IL-1{beta} reduces SDHB expression through upregulation of DNA methyltransferase 1 (DNMT1) and methylation of the SDHB promoter. Consequently, increased succinate levels impair oxidative phosphorylation and ATP synthesis, leading to reduced steroidogenesis. Together, we demonstrate that the IL-1{beta}-DNMT1-SDHB-succinate axis disrupts steroidogenesis. Our findings not only provide a mechanistic explanation for the adrenal dysfunction in severe inflammation but also a potential target for therapeutic intervention.

immunology↗

bioRxiv2020

Obesity-associated lipidomic remodeling of the adrenal gland indicates an important role of the FADS2-arachidonic acid axis in adrenocortical hormone production

ObjectiveAdrenocortical hormone levels increase in obesity, potentially contributing to development of obesity-associated pathologies. Here we explored whether lipidomic remodeling of the adrenal gland could mediate altered adrenocortical steroidogenesis during obesity. MethodsLipidomic analysis was performed in adrenal glands using shotgun mass spectrometry (MS), and steroid profiling of sera by liquid chromatography tandem mass spectrometry (LC-MS/MS) from lean and obese mice. Gene expression analysis was performed in adrenal glands and adrenocortical cell populations. The role of Fatty Acid Desaturase 2 (FADS2) and arachidonic acid on steroid hormone production was studied in primary adrenal gland cell cultures. ResultsAdrenal glands of obese mice displayed a distinct lipidomic profile, encompassing longer and more unsaturated storage lipids and phospholipids compared to adrenal glands of lean mice. Arachidonoyl acyl chains were abundant in the adrenal gland phospholipidome and increased upon obesity. This was accompanied by increased Fads2 expression, the rate-limiting enzyme of arachidonic acid synthesis, and enhanced plasma adrenocortical hormone levels. Inhibition of FADS2 in primary adrenal gland cell cultures abolished steroidogenesis, which was restored by arachidonic acid supplementation. ConclusionsOur data suggest that the FADS2 - arachidonic acid axis regulates adrenocortical hormone synthesis, while alterations in the content of arachidonoyl chains in the adrenal gland phopsholipidome could account for disturbed adrenocortical hormone production. HighlightsO_LIThe adrenal gland lipidome is remodeled in obesity. C_LIO_LIArachidonoyl groups are abundant in the adrenal gland phospholipidome and increase in obesity. C_LIO_LIFADS2 is highly expressed in the adrenal gland and its expression is further increased in obesity. C_LIO_LIFADS2 inhibition blunts adrenocortical steroidogenesis in primary adrenal gland cell cultures, while arachidonic acid supplementation restores it. C_LI

cell biology↗

bioRxiv2020

Modeling alcoholic liver disease in a human Liver-Chip

Fatty liver disease (FLD), is a major public health burden that affects up to 30% of people in Western countries and leads to progressive liver injury, comorbidities, and increased mortality. Key risk factors for developing FLD are obesity and alcohol consumption, both of which are growing in prevalence worldwide. There is an urgent need for human-relevant preclinical models to improve our understanding of FLD progression to steatohepatitis and for the development of sensitive noninvasive diagnostics and therapies. Alcohol-induced liver disease (ALD) represents an ideal case for modeling FDL as ethanol exposure is a comparatively simpler trigger for experimental induction of the pathology, as opposed to the complexity of modeling the diet- and life-style induced FLD. Further, despite their different root causes, the two pathologies share several common characteristics in disease progression and deterioration of liver function, highlighting the potential of an ALD microphysiological model for broad application in translational research. Here, we leverage our recently reported human Liver-Chip for toxicity applications, to expand the capabilities of the platform for broad application in translational research. We report the first in vitro modeling of ALD that uses human relevant blood alcohol concentrations (BAC) and affords multimodal profiling of clinically relevant endpoints. Our ALD Liver-Chip recapitulates established FLD markers in response to ethanol in a concentration-dependent manner, including lipid accumulation and oxidative stress. Importantly, we show that the ALD Liver-Chip supports the study of secondary insults common in patients with advanced ALD, such as high blood endotoxin levels due to alcohol-associated increased intestinal permeability and barrier dysfunction. Moreover, owing to new developments in the design, the ALD Liver-Chip enables the measurement of structural changes of the bile canaliculi (BC) network as a novel in vitro quantitative readout of alcoholic liver toxicity. In summary, we report the development of a human ALD Liver-Chip as a new platform for modeling the progression of alcohol-induced liver injury with direct translation to clinical research.

bioengineering↗