Biology subjects

Thomas, S. M.

Publications and source records attributed to Thomas, S. M..

10 recordsLinked to original sources

bioRxiv2026

Trans-Kingdom dsRNA Sensing: Aspergillus fumigatus Mycovirus Activates MDA5/MAVS Immunity and Limits Allergic Bronchopulmonary Aspergillosis

MDA5 is a cytosolic pattern-recognition receptor (PRR) that binds to double-stranded RNA (dsRNA) and subsequently interacts with the signaling adaptor protein MAVS to initiate the antiviral interferon (IFN) response. Our group previously demonstrated that MDA5 is essential for host resistance against the fungal pathogen, Aspergillus fumigatus. Although fungal dsRNA was sufficient to activate MDA5 signaling, the precise source of A. fumigatus dsRNA responsible for this MDA5-stimulating function remains unknown. Here, we demonstrate that the magnitude of the IFN-dependent antifungal response is A. fumigatus strain dependent. Unexpectedly, we found that A. fumigatus isolates infected with dsRNA mycoviruses triggered a more robust MAVS-dependent inflammatory response within alveolar macrophages. Furthermore, dsRNA mycovirus infection increased fungal susceptibility to antifungal killing without altering other A. fumigatus growth characteristics. Although dsRNA mycovirus infection did not alter virulence in an acute bronchopneumonia model of A. fumigatus infection, it significantly impaired virulence and improved disease parameters in a chronic model of allergic bronchopulmonary aspergillosis (ABPA). Collectively, these findings reveal a novel role for trans-kingdom interactions in driving the host antifungal IFN response and modulating virulence in chronic aspergillosis models.

immunology↗

bioRxiv2024

TGFβ primes alveolar-like macrophages to induce type I IFN following TLR2 activation

Alveolar macrophages (AMs) are key mediators of lung function and are potential targets for therapies during respiratory infections. TGF{beta} is an important regulator of AM differentiation and maintenance, but how TGF{beta} directly modulates the innate immune responses of AMs remains unclear. This shortcoming prevents effective targeting of AMs to improve lung function in health and disease. Here we leveraged an optimized ex vivo AM model system, fetal-liver derived alveolar-like macrophages (FLAMs), to dissect the role of TGF{beta} in AMs. Using transcriptional analysis, we first globally defined how TGF{beta} regulates gene expression of resting FLAMs. We found that TGF{beta} maintains the baseline metabolic state of AMs by driving lipid metabolism through oxidative phosphorylation and restricting inflammation. To better understand inflammatory regulation in FLAMs, we next directly tested how TGF{beta} alters the response to TLR2 agonists. While both TGF{beta} (+) and TGF{beta} (-) FLAMs robustly responded to TLR2 agonists, we found an unexpected activation of type I interferon (IFN) responses in FLAMs and primary AMs in a TGF{beta}-dependent manner. Surprisingly, mitochondrial antiviral signaling protein and the interferon regulator factors 3 and 7 were required for IFN production by TLR2 agonists and the IFN response was dependent on mitochondrial reactive oxygen species. Together, these data suggest that TGF{beta} modulates AM metabolic networks and innate immune signaling cascades to control inflammatory pathways in AMs.

immunology↗

bioRxiv2024

DCLK1-Mediated Regulation of Invadopodia Dynamics and Matrix Metalloproteinase Trafficking Drives Invasive Progression in Head and Neck Squamous Cell Carcinoma

Head and neck squamous cell carcinoma (HNSCC) is a major health concern due to its high mortality from poor treatment responses and locoregional tumor invasion into life sustaining structures in the head and neck. A deeper comprehension of HNSCC invasion mechanisms holds the potential to inform targeted therapies that may enhance patient survival. We previously reported that doublecortin like kinase 1 (DCLK1) regulates invasion of HNSCC cells. Here, we tested the hypothesis that DCLK1 regulates proteins within invadopodia to facilitate HNSCC invasion. Invadopodia are specialized subcellular protrusions secreting matrix metalloproteinases that degrade the extracellular matrix (ECM). Through a comprehensive proteome analysis comparing DCLK1 control and shDCLK1 conditions, our findings reveal that DCLK1 plays a pivotal role in regulating proteins that orchestrate cytoskeletal and ECM remodeling, contributing to cell invasion. Further, we demonstrate in TCGA datasets that DCLK1 levels correlate with increasing histological grade and lymph node metastasis. We identified higher expression of DCLK1 in the leading edge of HNSCC tissue. Knockdown of DCLK1 in HNSCC reduced the number of invadopodia, cell adhesion and colony formation. Using super resolution microscopy, we demonstrate localization of DCLK1 in invadopodia and colocalization with mature invadopodia markers TKS4, TKS5, cortactin and MT1-MMP. We carried out phosphoproteomics and validated using immunofluorescence and proximity ligation assays, the interaction between DCLK1 and motor protein KIF16B. Pharmacological inhibition or knockdown of DCLK1 reduced interaction with KIF16B, secretion of MMPs, and cell invasion. This research unveils a novel function of DCLK1 within invadopodia to regulate the trafficking of matrix degrading cargo. The work highlights the impact of targeting DCLK1 to inhibit locoregional invasion, a life-threatening attribute of HNSCC.

cancer biology↗

bioRxiv2023

Rapid lethality of mice lacking the phagocyte oxidase and Caspase1/11 following Mycobacterium tuberculosis infection.

Immune networks that control antimicrobial and inflammatory mechanisms have overlapping regulation and functions to ensure effective host responses. Genetic interaction studies of immune pathways that compare host responses in single and combined knockout backgrounds are a useful tool to identify new mechanisms of immune control during infection. For disease caused by pulmonary Mycobacterium tuberculosis infections, which currently lacks an effective vaccine, understanding genetic interactions between protective immune pathways may identify new therapeutic targets or disease-associated genes. Previous studies suggested a direct link between the activation of NLRP3-Caspase1 inflammasome and the NADPH-dependent phagocyte oxidase complex during Mtb infection. Loss of the phagocyte oxidase complex alone resulted in increased activation of Caspase1 and IL1{beta} production during Mtb infection, resulting in failed disease tolerance during the chronic stages of disease. To better understand this interaction, we generated mice lacking both Cybb, a key subunit of the phagocyte oxidase, and Caspase1/11. We found that ex vivo Mtb infection of Cybb-/-Caspase1/11-/- macrophages resulted in the expected loss of IL1{beta} secretion but an unexpected change in other inflammatory cytokines and bacterial control. Mtb infected Cybb-/-Caspase1/11-/- mice rapidly progressed to severe TB, succumbing within four weeks to disease characterized by high bacterial burden, increased inflammatory cytokines, and the recruitment of granulocytes that associated with Mtb in the lungs. These results uncover a key genetic interaction between the phagocyte oxidase complex and Caspase1/11 that controls protection against TB and highlight the need for a better understanding of the regulation of fundamental immune networks during Mtb infection.

immunology↗

bioRxiv2022

Doublecortin like kinase 1 is a target in squamous cell carcinoma

Doublecortin like kinase 1 (DCLK1) plays a crucial role in several cancers including colon and pancreatic adenocarcinomas. However, its role in squamous cell carcinoma (SCC) remains unknown. To this end, we examined DCLK1 expression in head and neck squamous cell carcinoma (HNSCC) and anal squamous cell carcinoma (ASCC). We found that DCLK1 is elevated in patient SCC tissue, which correlated with cancer progression and poorer overall survival. Furthermore, DCLK1 expression is significantly elevated in HPV negative cancer tissues, which are typically aggressive with poor responses to radiation therapy. To understand the role of DCLK1 in tumorigenesis, we used specific shRNA to suppress DCLK1 expression. This significantly reduced tumor growth, spheroid formation, and migration of HNSCC cancer cells. To further the translational relevance of our studies, we sought to identify a selective DCLK1 inhibitor. Current attempts to target DCLK1 using pharmacologic approaches have relied on non-specific suppression of DCLK1 kinase activity. Here, we demonstrate that DiFiD [3,5-bis (2,4-difluorobenzylidene)-4-piperidone] binds to DCLK1 with high selectivity. Moreover, DiFiD mediated suppression of DCLK1 led to G2/M arrest and apoptosis and significantly suppressed tumor growth of HNSCC xenografts and ASCC patient derived xenografts, supporting that DCLK1 is critical for SCC growth.

cancer biology↗

bioRxiv2021

RNA binding protein RBM3 augments kissing loop formation with lncRNAs to enhance translational control

It is becoming apparent that translational regulation involves the coordinated actions of RNA binding proteins (RBPs) and non-coding RNAs. For efficient translation, mRNA needs to be circularized, which is catalyzed by RNA binding proteins and translation factors. However, the role of lncRNAs in the process is not yet defined. We first performed RNA-seq and RNA- immunoprecipitation coupled-Seq and identified LSAMP-3 and Flii-1. Moreover, modeling studies suggest enhanced kissing loop interactions including of transcripts that encode angiogenesis and epithelial mesenchymal transition. While intestinal epithelial cell specific RBM3 transgenic mice showed increased LSAMP-3 and Flii-1, this was reduced in knockout mice. Also, RBM3 overexpression increased tumor xenograft growth, this was suppressed by knockdown of the lncRNAs. Also, knockdown of endogenous RBM3 reduced lncRNA levels and tumor xenograft growth. In addition, it reduced colitis-associated cancers. We propose that RBPs such as RBM3 mediate their function through regulatory lncRNAs that enable circularization to control translation.

cancer biology↗

bioRxiv2018

Y chromosomal noncoding RNA regulates autosomal gene expression via piRNAs in mouse testis

Majority of the genes expressed during spermatogenesis are autosomal. Mice with different deletions of Yq show sub-fertility, sterility and sperm abnormalities. The connection between Yq deletion and autosomal gene regulation is not well understood. We describe a novel mouse Yq-derived long noncoding RNA, Pirmy, which shows unprecedented number of splice variants in testis. Further, Pirmy transcript variants act as templates for several piRNAs. We identified ten differentially expressed autosome-encoded sperm proteins in mutant mice. Pirmy transcript variants have homology to 5/3UTRs of these deregulated autosomal genes. Thus, subfertility in Y-deleted mice appears to be a polygenic phenomenon that is partially regulated epistatically by the Y-chromosome. Our study provides novel insights into possible role of MSY-derived ncRNAs in male fertility and reproduction. Finally, sperm phenotypes from the Y-deleted mice seem to be similar to that reported in inter-specific male-sterile hybrids. Taken together, this study provides novel insights into possible role of Y-derived ncRNAs in male sterility and speciation.

genomics↗

bioRxiv2018

Broad-scale mercury bioaccumulation patterns in two freshwater sport fishes: testing the role of growth dilution in a warming climate

Sport fishes at the apex of aquatic food webs are indicators of mercury in the environment. However bioaccumulation of mercury in fish is a complex process that varies in space and time. Both large-scale climatic and environmental, as well as biological factors are drivers of these space-time variations. In this study, we avail a long-running monitoring program from Ontario, Canada to better understand spatiotemporal variations in fish mercury bioaccumulation. Focussing on two common large-bodied fishes (Walleye and Northern Pike), the data were first stratified by latitudinal zone (north, mid, and south) and eight temporal periods (between 1975 & 2015). A series of linear mixed-effects models (LMEMs) with latitudinal zone, time period, and their interactions as random effects were used to capture the spatial, temporal, and spatiotemporal variations in mercury bioaccumulation. The random slopes from the best-fitting LMEM were used to define bioaccumulation index and capture trends in space and time. Given the generally warming climate trend over the past 45 years, the role of growth dilution in modulating the bioaccumulation trends was also evaluated. The full model comprising of space, time and space-time interactions was the best-fit with interaction effects explaining most of the variation. Spatiotemporal trends showed overall similar patterns for both species. Growth dilution in conjunction with estimated rates of warming for different latitudinal zones failed to explain the spatiotemporal trends. Temporal trends showed contrasting bioaccumulation patterns-increasing in Northern Pike and decreasing in Walleye, suggesting temperature-driven growth dilution is more likely in latter. However, a space-for-time substitution revealed only a weak presence of growth dilution in Walleye, and it was not attributable to temperature differences. Overall, our study summarizes broad-scale variations in fish mercury and explores the role of growth dilution in shaping the observed patterns.

ecology↗

bioRxiv2017

A Comparative Study Of Endoderm Differentiation In Humans And Chimpanzees

There is substantial interest in the evolutionary forces that shaped the regulatory framework that is established in early human development. Progress in this area has been slow because it is difficult to obtain relevant biological samples. Inducible pluripotent stem cells (iPSCs) provide the ability to establish in vitro models of early human and non-human primate developmental stages. Using matched iPSC panels from humans and chimpanzees, we comparatively characterized gene regulatory changes through a four-day timecourse differentiation of iPSCs (day 0) into primary streak (day 1), endoderm progenitors (day 2), and definitive endoderm (day 3). As might be expected, we found that differentiation stage is the major driver of variation in gene expression levels, followed by species. We identified thousands of differentially expressed genes between humans and chimpanzees in each differentiation stage. Yet, when we considered gene-specific dynamic regulatory trajectories throughout the timecourse, we found that 75.. of genes, including nearly all known endoderm developmental markers, have similar trajectories in the two species. Interestingly, we observed a marked reduction of both intra-and inter-species variation in gene expression levels in primitive streak samples compared to the iPSCs, with a recovery of regulatory variation in endoderm progenitors. The reduction of variation in gene expression levels at a specific developmental stage, paired with overall high degree of conservation of temporal gene regulation, is consistent with the dynamics of developmental canalization. Overall, we conclude that endoderm development in iPSC-based models are highly conserved and canalized between humans and our closest evolutionary relative.

genomics↗

bioRxiv2016

Impact of regulatory variation across human iPSCs and differentiated cells

Induced pluripotent stem cells (iPSCs) are an essential tool for studying cellular differentiation and cell types that are otherwise difficult to access. We investigated the use of iPSCs and iPSC-derived cells to study the impact of genetic variation across different cell types and as models for studies of complex disease. We established a panel of iPSCs from 58 well-studied Yoruba lymphoblastoid cell lines (LCLs); 14 of these lines were further differentiated into cardiomyocytes. We characterized regulatory variation across individuals and cell types by measuring gene expression, chromatin accessibility and DNA methylation. Regulatory variation between individuals is lower in iPSCs than in the differentiated cell types, consistent with the intuition that developmental processes are generally canalized. While most cell type-specific regulatory quantitative trait loci (QTLs) lie in chromatin that is open only in the affected cell types, we found that 20% of cell type-specific QTLs are in shared open chromatin. Finally, we developed a deep neural network to predict open chromatin regions from DNA sequence alone and were able to use the sequences of segregating haplotypes to predict the effects of common SNPs on cell type-specific chromatin accessibility.

genomics↗