Biology subjects

Lee, J. C.

Publications and source records attributed to Lee, J. C..

36 records · Page 2Linked to original sources

bioRxiv2023

The lung employs an intrinsic surfactant-mediated inflammatory response for viral defense

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) causes an acute respiratory distress syndrome (ARDS) that resembles surfactant deficient RDS. Using a novel multi-cell type, human induced pluripotent stem cell (hiPSC)-derived lung organoid (LO) system, validated against primary lung cells, we found that inflammatory cytokine/chemokine production and interferon (IFN) responses are dynamically regulated autonomously within the lung following SARS-CoV-2 infection, an intrinsic defense mechanism mediated by surfactant proteins (SP). Single cell RNA sequencing revealed broad infectability of most lung cell types through canonical (ACE2) and non-canonical (endocytotic) viral entry routes. SARS-CoV-2 triggers rapid apoptosis, impairing viral dissemination. In the absence of surfactant protein B (SP-B), resistance to infection was impaired and cytokine/chemokine production and IFN responses were modulated. Exogenous surfactant, recombinant SP-B, or genomic correction of the SP-B deletion restored resistance to SARS-CoV-2 and improved viability.

immunology↗

bioRxiv2022

EVIDENCE FOR EMPATHY IN PAIR BONDING POISON FROGS

More than a century ago, Charles Darwin hypothesized that the empathy-like phenotype is a phylogenetically widespread phenomenon. This idea remains contentious, due to the challenges of empirically examining emotions, and few investigations among non-mammalian vertebrates. We provide support for Darwins hypothesis by discovering partial evidence for the most ancestral form of empathy, emotional contagion (i.e., matching another individuals emotional state), in the pair bonding mimetic poison frog, Ranitomeya imitator. We found that male corticosterone, a physiological biomarker of stress, positively correlates with female partners in experimental and semi-natural conditions. This does not appear to coincide with behavioral state-matching. However, it is specific to female partners relative to familiar female non-partners, and is independent of effects that commonly confound studies on emotional contagion. Furthermore, this physiological state-matching is irrespective of partnership longevity or lifetime reproductive output. These results physiologically indicate socially selective emotional contagion in a monogamous amphibian, and paradigms that elicit coinciding neural and behavioral indicators and morphogenic co-variation are needed for further corroboration. Further studies on ancestral forms of empathy in non-mammalian vertebrates are warranted.

animal behavior and cognition↗

bioRxiv2021

Beta-Catenin Limits Osteogenesis on Regenerative Materials in a Stiffness-Dependent Manner

Targeted refinement of regenerative materials requires mechanistic understanding of cell-material interactions. The nanoparticulate mineralized collagen glycosaminoglycan (MC-GAG) scaffold is a porous biomaterial that promotes regenerative healing of calvaria defects in vivo without addition of exogenous growth factors or progenitor cells, suggesting its potential as an off-the-shelf implant for reconstructing skull defects. In this work, we evaluate the relationship between material stiffness, a tunable MC-GAG property, and activation of the canonical Wnt (cWnt) signaling pathway. Primary human bone marrow-derived mesenchymal stem cells (hMSCs) were differentiated on two MC-GAG scaffolds varying by stiffness (non- crosslinked, NX-MC, 0.3 kPa vs. conventionally crosslinked, MC, 3.9 kPa). hMSCs exhibited increased expression of activated {beta}-catenin, the major cWnt intracellular mediator, and the mechanosensitive YAP protein with near complete subcellular colocalization in stiffer MC scaffolds. Small molecule Wnt pathway inhibitors reduced activated {beta}-catenin and YAP protein quantities and colocalization, osteogenic differentiation, and mineralization on MC, with no effects on NX-MC. Concomitantly, Wnt inhibitors increased BMP4 and phosphorylated Smad1/5 (p-Smad1/5) expression on MC, but not NX-MC. Unlike non-specific Wnt pathway downregulation, isolated canonical Wnt inhibition with {beta}-catenin knockdown increased osteogenic gene expression and mineralization specifically on the stiffer MC. {beta}-catenin knockdown also increased p-Smad1/5, Runx2, and BMP4 expression only on the stiffer MC material. Our data indicates stiffness-induced activation of the Wnt and mechanotransduction pathways promotes osteogenesis in MC-GAG scaffolds. However, activated {beta}-catenin is a limiting agent and may serve as a useful target or readout for optimal modulation of stiffness in skeletal regenerative materials. One Sentence Summary{beta}-Catenin limits stiffness-induced osteogenenic differentiation on nanoparticulate mineralized collagen glycosaminoglycan materials

bioengineering↗

bioRxiv2021

A Functional Menadione Biosynthesis Pathway is Required for Capsule Production by Staphylococcus aureus.

Staphylococcus aureus is a major human pathogen that utilises a wide array of pathogenic and immune evasion strategies to cause disease. One immune evasion strategy, common to many bacterial pathogens, is the ability of S. aureus to produce a capsule that protects the bacteria from several aspects of the human immune system. To identify novel regulators of capsule production by S. aureus we applied a genome wide association study (GWAS) to a collection of 300 bacteraemia isolates that represent the two major MRSA clones in UK and Irish hospitals: CC22 and CC30. One of the loci associated with capsule production, the menD gene, encodes an enzyme critical to the biosynthesis of menadione. Mutations in this gene that result in menadione auxotrophy induce the slow growing small-colony variant (SCV) form of S. aureus often associated with chronic infections due to their increased resistance to antibiotics and ability to survive inside phagocytes. Utilising such an SCV we functionally verified this association between menD and capsule production. Although the clinical isolates with polymorphisms in the menD gene in our collections had no apparent growth defects, they were more resistant to gentamicin when compared to those with the wild-type menD gene. Our work suggests that menadione plays a critical role in the production of the S. aureus capsule, and that amongst clinical isolates polymorphisms exist in the menD gene that confer the characteristic increased gentamicin resistance, but not the major growth defect associated with SCV phenotype.

microbiology↗

bioRxiv2021

Multiomic Characterization of Stage I Lung Adenocarcinoma Reveals Distinct Genetic and Immunologic Features of Recurrent Disease

BackgroundRecurrence after surgery for early-stage lung cancer is common, occurring between 30-50% of the time. Despite the popularization of prognostic gene signatures in early-stage lung cancer that allow us to better predict which patients may recur, why patients recur after surgery remains unclear. MethodsUsing a large cohort of lung adenocarcinoma patients with complete genetic, genomic, epigenetic and clinical profiling, a recurrence classifier was developed which identifies patients at highest risk of recurrence. The genetic, genomic, and epigenetic profiles of stage I patients with low-vs. high-risk of recurrence were compared. To characterize the tumor immune microenvironment of recurrent stage I tumors, single cell RNA-seq was performed on fresh tissue samples undergoing lung adenocarcinoma resection at UCSF to identify unique immune population markers and applied to the large stage I lung adenocarcinoma cohort using digital cytometry. ResultsRecurrence high-risk stage I lung adenocarcinomas demonstrated a higher mutation burden than low-risk tumors, however, none of the known canonical lung cancer driver mutations were more prevalent in high-risk tumors. Transcriptomic analysis revealed widespread activation of known cancer and cell cycle pathways with simultaneous downregulation of immune response pathways including antigen presentation and Th1/Th2 activation. Tumors at high-risk of recurrence displayed depleted adaptive immune populations, and depletion of adaptive immune populations was independently prognostic of recurrence in stage I lung adenocarcinomas. ConclusionRecurrent stage I lung adenocarcinomas display distinct features of genomic and genetic instability including increased tumor mutation burden, neoantigen load, activation of numerous mitotic and cell cycle genes, and decreased genome-wide methylation burden. Relative depletion of infiltrating adaptive immune populations may allow these tumors to escape immunosurveillance and recur after surgery.

cancer biology↗

bioRxiv2021

Punishment insensitivity in humans is due to failures in instrumental contingency learning

Punishment maximises the probability of our individual survival by reducing behaviours that cause us harm, and also sustains trust and fairness in groups essential for social cohesion. However, some individuals are more sensitive to punishment than others and these differences in punishment sensitivity have been linked to a variety of decision-making deficits and psychopathologies. The mechanisms for why individuals differ in punishment sensitivity are poorly understood, although recent studies of conditioned punishment in rodents highlight a key role for punishment contingency detection (Jean-Richard-dit-Bressel et al., 2019). Here we applied a novel "Planets & Pirates" conditioned punishment task in humans, allowing us to identify the mechanisms for why individuals differ in their sensitivity to punishment. We show that punishment sensitivity is bimodally distributed in a large sample of normal participants. Sensitive and insensitive individuals equally liked reward and showed similar rates of reward-seeking. They also equally disliked punishment and did not differ in their valuation of cues that signalled punishment. However, sensitive and insensitive individuals differed profoundly in their capacity to detect and learn volitional control over aversive outcomes. Punishment insensitive individuals did not learn the instrumental contingencies, so they could not withhold behaviour that caused punishment and could not generate appropriately selective behaviours to prevent impending punishment. These differences in punishment sensitivity could not be explained by individual differences in behavioural inhibition, impulsivity, or anxiety. This bimodal punishment sensitivity and these deficits in instrumental contingency learning are identical to those dictating punishment sensitivity in non-human animals, suggesting that they are general properties of aversive learning and decision making.

neuroscience↗

bioRxiv2021

Genetically Engineered Strains for Combinatorial Cardiovascular Optobiology

Optogenetic effectors and sensors provide a novel real-time window into complex physiological processes, enabling determination of molecular signaling processes within functioning cellular networks. However, the combination of these optical tools in mice is made practical by construction of genetic lines that are optically compatible and genetically tractable. We present a new toolbox of 21 mouse lines with lineage -specific expression of optogenetic effectors and sensors for direct biallelic combination, avoiding the multiallelic requirement of Cre recombinase -mediated DNA recombination, focusing on models relevant for cardiovascular biology. Optogenetic effectors (11 lines) or Ca2+ sensors (10 lines) were selectively expressed in cardiac pacemaker cells, cardiomyocytes, vascular endothelial and smooth muscle cells, alveolar epithelial cells, lymphocytes, glia, and other cell types. Optogenetic effector and sensor function was demonstrated in numerous tissues. Arterial/arteriolar tone was modulated by optical activation of the second messengers InsP3 (opto1AR) and cAMP (opto{beta}2AR), or Ca2+- permeant membrane channels (CatCh2) in smooth muscle (Acta2) and endothelium (Cdh5). Cardiac activation was separately controlled through activation of nodal/conducting cells or cardiac myocytes. We demonstrate combined effector and sensor function in biallelic mouse crosses: optical cardiac pacing and simultaneous cardiomyocyte Ca2+ imaging in HCN4BAC-CatCh2/MHC-GCaMP8 crosses. These experiments highlight the potential of these mice to explore cellular signaling in vivo, in complex tissue networks.

physiology↗

bioRxiv2021

Trans-channel fluorescence learning improves high-content screening for Alzheimer's disease therapeutics

In microscopy-based drug screens, fluorescent markers carry critical information on how compounds affect different biological processes. However, practical considerations may hinder the use of certain fluorescent markers. Here, we present a deep learning method for overcoming this limitation. We accurately generated predicted fluorescent signals from other related markers and validated this new machine learning (ML) method on two biologically distinct datasets. We used the ML method to improve the selection of biologically active compounds for Alzheimers disease (AD) from high-content high-throughput screening (HCS). The ML method identified novel compounds that effectively blocked tau aggregation, which would have been missed by traditional screening approaches unguided by ML. The method improved triaging efficiency of compound rankings over conventional rankings by raw image channels. We reproduced this ML pipeline on a biologically independent cancer-based dataset, demonstrating its generalizability. The approach is disease-agnostic and applicable across diverse fluorescence microscopy datasets.

pharmacology and toxicology↗

bioRxiv2020

In situ differentiation of iridophore crystallotypes underlies zebrafish stripe patterning

Skin color patterns are ubiquitous in nature, evolve rapidly, and impact social behavior1, predator avoidance2, and protection from ultraviolet irradiation3. A leading model system for vertebrate skin patterning is the zebrafish4-7; its alternating blue stripes and yellow interstripes depend on guanine crystal-containing cells called iridophores that reflect light. It was suggested that the zebrafishs alternating color pattern arises from a single type of iridophore migrating differentially to stripes and interstripes7-9. When we tracked iridophores, however, we found they did not migrate between stripes and interstripes but instead differentiated and proliferated in place based on their micro-environment. RNA seq analysis further revealed stripe and interstripe iridophores had different transcriptomic states, while cryogenic scanning electron microscopy and micro-X-ray diffraction showed they had different guanine crystal organizations and responsiveness to norepinephrine, all indicating that stripe and interstripe iridophores are different cell types. Based on these results, we present a new model of skin patterning in zebrafish in which distinct iridophore crystallotypes containing specialized, physiologically responsive, subcellular organelles arise in stripe and interstripe zones by in situ differentiation. In this model, pattern phenotype depends not only on interactions among pigment cells that affect their arrangements, but also on factors that specify subcellular organization and physiological responsiveness of specialized organelles.

developmental biology↗

bioRxiv2020

Stiffness of Nanoparticulate Mineralized Collagen Scaffolds Triggers Osteogenesis via Mechanotransduction and Canonical Wnt Signaling

The ability of the extracellular matrix (ECM) to instruct progenitor cell differentiation has generated excitement for the development of materials-based regenerative solutions. We previously described a nanoparticulate mineralized collagen glycosaminoglycan (MC-GAG) material capable of inducing in vivo skull regeneration approaching 60% of the biomechanical properties of native calvarium without exogenous growth factors or ex vivo progenitor cell-priming, suggesting promise as a first-generation material for skull regeneration. Here, we evaluated the contribution of titrating stiffness to osteogenicity by comparing non-crosslinked (NX-MC) and crosslinked (MC) forms of MC-GAG. While both materials were osteogenic, MC demonstrated an increased expression of osteogenic markers and mineralization compared to NX-MC. Both materials were capable of autogenously activating the canonical bone morphogenetic protein receptor (BMPR) signaling pathway with phosphorylation of Smad1/5 (small mothers against decapentaplegic-1/5). However, unlike NX-MC, hMSCs cultured on MC demonstrated significant elevations in the major mechanotransduction mediators YAP (Yes-associated protein) and TAZ (transcriptional coactivator with PDZ-binding motif) expression coincident with {beta}-catenin activation in the canonical Wnt signaling pathway. Inhibition of YAP/TAZ activation reduced osteogenic marker expression, mineralization, and {beta}-catenin activation in MC with a much lesser of an effect on NX-MC. YAP/TAZ inhibition also resulted in a reciprocal increase in Smad1/5 phosphorylation as well as BMP2 expression. Our results indicate that increasing MC-GAG stiffness induces osteogenic differentiation via the mechanotransduction mediators YAP/TAZ and the canonical Wnt signaling pathway, whereas the canonical BMPR signaling pathway is activated in a manner independent of mechanical cues.

bioengineering↗

bioRxiv2020

Resolving mechanisms of immune-mediated disease in primary CD4 T cells

Deriving mechanisms of immune-mediated disease from GWAS data remains a formidable challenge, with attempts to identify causal variants being frequently hampered by linkage disequilibrium. To determine whether causal variants could be identified via their functional effects, we adapted a massively-parallel reporter assay for use in primary CD4 T-cells, key effectors of many immune-mediated diseases. Using the results to guide further study, we provide a generalisable framework for resolving disease mechanisms from non-coding associations - illustrated by a locus linked to 6 immune-mediated diseases, where the lead functional variant causally disrupts a super-enhancer within an NF-{kappa}B-driven regulatory circuit, triggering unrestrained T-cell activation.

genetics↗

bioRxiv2020

Characterisation of the genetic architecture of immune-mediated disease through informed dimension reduction

Integration of genome-wide association study (GWAS) data has been used to generate new hypotheses of biological mechanism, aetiological relationships between traits, or test causality of one factor for another. However, such approaches have typically been limited to pairwise comparisons of traits. We propose a generally applicable method, that exploits ideas from Bayesian genetic fine mapping to define a "lens" that focuses relevant variants before dimension reduction of a set of related GWAS summary statistics. We applied this technique to immune-mediated diseases, deriving 13 components which summarise the multidimensional patterns of genetic risk. Projection of independent datasets demonstrated the specificity and accuracy of our reduced dimension basis, enabled us to functionally characterise individual components, identify disease-discriminating components and suggest novel associations in rare diseases where classical GWAS approaches are challenging. Our approach summarises the genetic architectures underlying any range of aetiologically-related traits in fewer dimensions, facilitating more nuanced multidimensional comparative analyses.

genetics↗

bioRxiv2019

The Firre locus produces a trans-acting RNA molecule that functions in hematopoiesis

RNA has been classically known to play central roles in biology, including maintaining telomeres1, protein synthesis2, and in sex chromosome compensation in certain species3,4. At the center of these important biological systems are noncoding RNAs. While thousands of long noncoding RNAs (lncRNAs) have been identified in mammalian genomes5-8, attributing RNA-based roles to lncRNA loci requires an assessment of whether the observed effect could be due to DNA regulatory elements, the act of transcription, or the lncRNA transcript. Here, we use the syntenically conserved lncRNA locus, Functional intergenic repeating RNA element (Firre), that is located on the X chromosome as a model to discriminate between DNA- and RNA-mediated effects in vivo. To this end, we generated genetically defined loss-of-function, gain-of-function, and rescue mouse models for Firre and provide genetic evidence that the Firre locus produces a trans-acting RNA. We report that: (i) Firre mutant mice have cell-specific defects during hematopoiesis and changes in gene expression that can be rescued by induction of Firre RNA from a transgene in the Firre knockout background, (ii) mice overexpressing Firre from a transgene exhibit increased levels of pro-inflammatory cytokines and impaired survival upon exposure to lipopolysaccharide, and (iii) deletion of the Firre locus did not result in changes in local gene expression on the X chromosome in 9 different biological contexts, suggesting that Firre does not function by cis-acting RNA or DNA elements. Together, our results provide genetic evidence that the Firre locus produces a trans-acting lncRNA that has physiological roles in hematopoiesis and immune function.

immunology↗

bioRxiv2019

Revisiting the regulation of the capsular polysaccharide biosynthesis gene cluster in Staphylococcus aureus

In Staphylococcus aureus, the capsular polysaccharide (CP) protects against phagocytosis, but also hinders adherence to endothelial cells and matrix proteins. Its biosynthesis is tightly controlled resulting in a heterogeneous phenotype within a population and CP being mainly detectable in non-growing cells. Capsular biosynthesis genes are encoded by a conserved capA-P operon whose expression is driven by an upstream promoter element (Pcap) in front of capA. The organization of Pcap is poorly understood, as is the interplay of different regulators that influence the early-Off/late-Heterogeneous cap transcription pattern. Here, we demonstrate that Pcap contains a main SigB-dependent promoter. The SigB consensus motif overlaps with a previously described inverted repeat that is crucial for cap expression. The essentiality of the inverted repeat is derived from this region acting as a SigB binding site rather than as an operator site for the proposed cap activators RbsR and MsaB. Furthermore, Pcap contains an extensive upstream region harboring a weak SigA-dependent promoter and binding sites for the cap repressors SaeR, CodY and Rot. We show that heterogeneous CP synthesis is determined by the combination of SigB activity and repressor binding to the upstream region. The direct SigB dependency and the upstream repressors are also sufficient to explain the temporal gene expression pattern at the transcriptional level. However, CP synthesis remains growth phase-dependent even when capA transcription is rendered constitutive, suggesting additional post-transcriptional regulatory circuits. Thus, the interference of multiple repressors with SigB-dependent promoter activity as well as post-transcriptional mechanisms ensure the appropriate regulation of CP synthesis.\n\nImportanceThe majority of bacterial pathogens produce an array of polysaccharides on their surface which are important virulence factors and thus serve as attractive vaccine candidates. However, the synthesis and assembly of these structures is highly variable and tightly regulated at various levels. In the human pathogen Staphylococcus aureus, the synthesis of the capsular polysaccharide (CP) is dependent on a complex regulatory network which ensures that CP is produced only in a fraction of stationary phase cells. Here, we determined main regulators that drive the peculiar CP expression pattern. We found that the interplay of the transcriptional repressors Sae, CodY and Rot with the alternative Sigma factor B is responsible for early-Off/late-Heterogeneous expression at the transcriptional level. The data also implicates post-transcriptional mechanisms that may act to avoid conflict in precursor usage by machineries involved in either synthesis of CP or other glycopolymers in growing bacterial cells.

microbiology↗

bioRxiv2019

A blood-based prognostic biomarker in inflammatory bowel disease

ObjectiveWe have previously described a prognostic transcriptional signature in CD8 T cells that separates inflammatory bowel disease (IBD) patients into 2 phenotypically-distinct subgroups, which we termed IBD1 and IBD2. Here we sought to develop a blood-based prognostic test that could identify these patient subgroups without the need for cell separation, and thus be suitable for routine clinical use in Crohns disease (CD) and ulcerative colitis (UC). DesignPatients with active IBD were recruited before treatment. Transcriptomic analyses were performed on purified CD8 T cells and/or whole blood. Detailed phenotype information was collected prospectively. IBD1/IBD2 patient subgroups were identified by consensus clustering of CD8 T cell transcriptomes. In a training cohort, statistical (machine) learning was used to identify groups of genes ("classifiers") whose differential expression in whole blood re-created the IBD1/IBD2 patient subgroups. Genes from the best classifiers were qPCR-optimised, and further statistical learning was applied to the qPCR dataset to identify the optimal classifier, which was locked-down for further testing. Independent validation was sought in separate cohorts of CD (n=66) and UC patients (n=57). ResultsIn both independent validation cohorts, a 17-gene qPCR-based classifier stratified patients into two distinct subgroups. Irrespective of the underlying diagnosis, IBDhi patients (analogous to the poor prognosis IBD1 subgroup) experienced significantly more aggressive disease than IBDlo patients (analogous to IBD2), with earlier need for treatment escalation and more escalations over time. ConclusionThis is the first validated prognostic biomarker that can predict prognosis in newly-diagnosed IBD patients, and represents a step towards personalised therapy. What is already known about this subject?The course of CD and UC varies considerably between patients, but reliable prognostic markers are not available in clinical practice. This hinders disease management because treatment approaches that would be optimal for patients with indolent disease - characterised by infrequent flare-ups that can be readily controlled by first-line therapy - will inevitably undertreat those with progressive disease. Conversely, strategies that would appropriately control frequently-relapsing, progressive disease will expose patients with more quiescent disease to the risks and side effects of unnecessary treatment. We have previously described a CD8 T cell gene expression signature that corresponds to differences in T cell exhaustion, is detectable during active untreated disease (including at diagnosis), and predicts disease course in both UC and CD. However, the need for cell separation and microarray-based gene expression analysis would make this difficult to translate to clinical practice. What are the new findings?We have developed, optimised, and independently validated a whole blood qPCR-based classifier - designed to identify the IBD1 and IBD2 patient subgroups - that can reliably predict prognosis in patients with CD or UC from diagnosis without the need for cell separation. We also present a detailed phenotypic update on the disease course experienced by patients in either the IBD1/IBDhi or IBD2/IBDlo subgroups, incorporating both expanded patient cohorts and substantially longer follow-up. This affords new insights into the spectrum of therapies that are differentially required in these patient subgroups, and reinforces their association with disease prognosis. How might it impact on clinical practice in the foreseeable future?The qPCR-based classifier has performance characteristics that compare favourably to prognostic biomarkers currently in use in oncology, and should be sufficient to guide therapy from diagnosis in patients with CD or UC. This represents an important step towards personalised therapy in inflammatory bowel disease.

genomics↗

bioRxiv2018

High-throughput functional analysis of lncRNA core promoters elucidates rules governing tissue-specificity

Transcription initiates at both coding and non-coding genomic elements, including mRNA and long non-coding RNA (lncRNA) core promoters and enhancer RNAs (eRNAs). However, each class has different expression profiles with lncRNAs and eRNAs being the most tissue-specific. How these complex differences in expression profiles and tissue-specificities are encoded in a single DNA sequence, however, remains unresolved. Here, we address this question using computational approaches and massively parallel reporter assays (MPRA) surveying hundreds of promoters and enhancers. We find that both divergent lncRNA and mRNA core promoters have higher capacities to drive transcription than non-divergent lncRNA and mRNA core promoters, respectively. Conversely, lincRNAs and eRNAs have lower capacities to drive transcription and are more tissue-specific than divergent genes. This higher tissue-specificity is strongly associated with having less complex TF motif profiles at the core promoter. We experimentally validated these findings by testing both engineered single-nucleotide deletions and human single-nucleotide polymorphisms (SNPs) in MPRA. In both cases, we observe that single nucleotides associated with many motifs are important drivers of promoter activity. Thus, we suggest that high TF motif density serves as a robust mechanism to increase promoter activity at the expense of tissue-specificity. Moreover, we find that 22% of common SNPs in core promoter regions have significant regulatory effects. Collectively, our findings show that high TF motif density provides redundancy and increases promoter activity at the expense of tissue specificity, suggesting that specificity of expression may be regulated by simplicity of motif usage.

genomics↗

bioRxiv2018

Adjustment for index event bias in genome-wide association studies of subsequent events

Following numerous genome-wide association studies of disease susceptibility, there is increasing interest in genetic associations with disease prognosis, survival or other subsequent events. Such associations are vulnerable to index event bias, by which selection of subjects according to their disease status creates biased associations if common causes of incidence and prognosis are not accounted for. We propose a novel adjustment for index event bias using the residuals from the regression of genetic effects on prognosis on genetic effects on incidence. Our approach eliminates this bias when direct genetic effects on incidence and prognosis are independent, and otherwise reduces bias in realistic situations. In a study of idiopathic pulmonary fibrosis, we resolved a paradoxical association of the strong susceptibility gene MUC5B with increased survival, identifying instead a significant association with decreased survival. In re-analysis of a study of Crohns disease prognosis, four regions remained associated at genome-wide significance albeit with increased P-values.

epidemiology↗

bioRxiv2018

The biogenesis of extracellular vesicles from Staphylococcus aureus and their application as a novel vaccine platform

Gram-positive bacteria secrete extracellular vesicles (EVs) that package diverse bacterial antigens and play key roles in bacterial pathogenesis. However, the mechanisms underlying EV production in Gram-positive bacteria are poorly understood. We purified and characterized EVs from a community-associated methicillin-resistant Staphylococcus aureus isolate (USA300) and investigated mechanisms underlying EV production. Native EVs contained 165 proteins, including cytosolic, surface, and secreted proteins, autolysins, and numerous cytolysins. Staphylococcal alpha-type phenol-soluble modulins (surfactant-like peptides) promoted EV biogenesis, presumably by acting at the cytoplasmic membrane, whereas peptidoglycan crosslinking and autolysin activity were found to increase EV production by altering the permeability of the staphylococcal cell wall. To address the immunogenicity of EVs, we created engineered EVs (eng-EVs) by expressing detoxified proteins HlaH35L and LukE in EVs generated from a nontoxic S. aureus {Delta}agr{Delta}spa mutant. Eng-EVs exhibited no cytotoxicity in vitro, and mice immunized with the eng-EVs produced toxin-neutralizing antibodies and showed reduced lethality in a mouse sepsis model. Our study reveals novel mechanisms underlying S. aureus EV production and highlights the usefulness of EVs as a novel S. aureus vaccine platform.

microbiology↗