Biology subjects

Lafyatis, R.

Publications and source records attributed to Lafyatis, R..

9 recordsLinked to original sources

bioRxiv2025

Suppression of interferon signaling via small molecule modulation of TFAM

The mitochondrial transcription factor A (TFAM) is essential for mitochondrial genome maintenance. It binds to mitochondrial DNA (mtDNA) and determines the abundance, packaging and stability of the mitochondrial genome. Because its function is tightly associated with mtDNA, TFAM has a protective role in mitochondrial diseases and supportive studies demonstrate reversal of disease phenotypes by TFAM overexpression. In addition, TFAM deficiency has been shown to cause release of mtDNA into the cytosol and activation of the cGAS/STING innate immune response pathway. As such, TFAM presents as a unique target for therapeutic intervention, but limited efforts for activators have been reported. Herein we disclose novel TFAM small molecule modulators with sub-micromolar activity. Our results demonstrate that these compounds result in increase of TFAM protein levels and mtDNA copy number. This results in inhibition of a mtDNA stress-mediated inflammatory response by preventing mtDNA escape into the cytosol. Furthermore, we see beneficial effects in cellular disease models in which boosting TFAM activity has been advanced as a disease modifying strategy including improved energetics in MELAS cybrid cells and a decrease of fibrotic markers in Systemic Sclerosis (SSc) fibroblasts. These results highlight the therapeutic potential of using small molecule TFAM activators in indications characterized by mitochondrial dysfunction.

cell biology↗

bioRxiv2024

SenSet, a novel human lung senescence cell gene signature, identifies cell-specific senescence mechanisms

Cellular senescence is a major hallmark of aging. Senescence is defined as an irreversible growth arrest observed when cells are exposed to a variety of stressors including DNA damage, oxidative stress, or nutrient deprivation. While senescence is a well-established driver of aging and age-related diseases, it is a highly heterogeneous process with significant variations across organisms, tissues, and cell types. The relatively low abundance of senescence in healthy aged tissues represents a major challenge to studying senescence in a given organ, including the human lung. To overcome this limitation, we developed a Positive-Unlabeled (PU) learning framework to generate a comprehensive senescence marker gene list in human lungs (termed SenSet) using the largest publicly available single-cell lung dataset, the Human Lung Cell Atlas (HLCA). We validated SenSet in a highly complex ex vivo human 3D lung tissue culture model subjected to the senescence inducers bleomycin, doxorubicin, or irradiation, and established its sensitivity and accuracy in characterizing senescence. Using SenSet, we identified and validated cell-type specific senescence signatures in distinct lung cell populations upon aging and environmental exposures. Our study presents the first comprehensive analysis of senescent cells in the healthy aging lung and uncovers cell-specific gene signatures of senescence, presenting fundamental implications for our understanding of major lung diseases, including cancer, fibrosis, chronic obstructive pulmonary disease, or asthma.

genomics↗

bioRxiv2024

Altered AP-1, RUNX, and EGR chromatin dynamics drive fibrotic lung disease

Pulmonary fibrosis, including systemic sclerosis-associated interstitial lung disease (SSc-ILD), involves myofibroblasts and SPP1hi macrophages as drivers of fibrosis. Single-cell RNA sequencing has delineated fibroblast and macrophages transcriptomes, but limited insight into transcriptional control of profibrotic gene programs. To address this challenge, we analyzed multiomic snATAC/snRNA-seq on explanted SSc-ILD and donor control lungs. The neural network tool ChromBPNet inferred increased TF binding at single base pair resolution to profibrotic genes, including CTHRC1 and ADAM12, in fibroblasts and SPP1 and CCL18 in macrophages. The novel algorithm HALO confirmed AP-1, RUNX, and EGR TF activity controlling profibrotic gene programs and established TF-regulatory element-gene networks. This TF action atlas provides comprehensive insights into the transcriptional regulation of fibroblasts and macrophages in healthy and fibrotic human lungs.

systems biology↗

bioRxiv2024

Genetic regulation and targeted reversal of lysosomal dysfunction and inflammatory sterol metabolism in pulmonary arterial hypertension

Vascular inflammation critically regulates endothelial cell (EC) pathophenotypes, particularly in pulmonary arterial hypertension (PAH). Dysregulation of lysosomal activity and cholesterol metabolism have known inflammatory roles in disease, but their relevance to PAH is unclear. In human pulmonary arterial ECs and in PAH, we found that inflammatory cytokine induction of the nuclear receptor coactivator 7 (NCOA7) both preserved lysosomal acidification and served as a homeostatic brake to constrain EC immunoactivation. Conversely, NCOA7 deficiency promoted lysosomal dysfunction and proinflammatory oxysterol/bile acid generation that, in turn, contributed to EC pathophenotypes. In vivo, mice deficient for Ncoa7 or exposed to the inflammatory bile acid 7-hydroxy-3-oxo-4-cholestenoic acid (7HOCA) displayed worsened PAH. Emphasizing this mechanism in human PAH, an unbiased, metabolome-wide association study (N=2,756) identified a plasma signature of the same NCOA7-dependent oxysterols/bile acids associated with PAH mortality (P<1.1x10-6). Supporting a genetic predisposition to NCOA7 deficiency, in genome-edited, stem cell-derived ECs, the common variant intronic SNP rs11154337 in NCOA7 regulated NCOA7 expression, lysosomal activity, oxysterol/bile acid production, and EC immunoactivation. Correspondingly, SNP rs11154337 was associated with PAH severity via six-minute walk distance and mortality in discovery (N=93, P=0.0250; HR=0.44, 95% CI [0.21-0.90]) and validation (N=630, P=2x10-4; HR=0.49, 95% CI [0.34-0.71]) cohorts. Finally, utilizing computational modeling of small molecule binding to NCOA7, we predicted and synthesized a novel activator of NCOA7 that prevented EC immunoactivation and reversed indices of rodent PAH. In summary, we have established a genetic and metabolic paradigm and a novel therapeutic agent that links lysosomal biology as well as oxysterol and bile acid processes to EC inflammation and PAH pathobiology. This paradigm carries broad implications for diagnostic and therapeutic development in PAH and in other conditions dependent upon acquired and innate immune regulation of vascular disease. One Sentence SummaryPulmonary arterial hypertension pathophenotypes arise from allele-specific NCOA7 regulation of lysosome function and inflammatory oxysterol generation, as demonstrated by genomic and metabolomic association studies coupled with genetic and pharmacologic mechanistic evidence. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=112 SRC="FIGDIR/small/582142v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@896f6forg.highwire.dtl.DTLVardef@f31c2eorg.highwire.dtl.DTLVardef@bf46dcorg.highwire.dtl.DTLVardef@1c55686_HPS_FORMAT_FIGEXP M_FIG Graphical abstract. C_FIG

physiology↗

bioRxiv2021

An instance-specific causal framework for learning intercellular communication networks that define microenvironments of individual tumors

Cells within a tumor microenvironment (TME) dynamically communicate and influence each others cellular states through an intercellular communication network (ICN). In cancers, intercellular communications underlie immune evasion mechanisms of individual tumors. We developed an individualized causal analysis framework for discovering tumor-specific ICNs. Using head and neck squamous cell carcinoma (HNSCC) tumors as a testbed, we first mined single-cell RNA-sequencing data to discover gene expression modules (GEMs) that reflect the states of transcriptomic processes within tumor and stromal single cells. By deconvoluting bulk transcriptomes of HNSCC tumors profiled by The Cancer Genome Atlas (TCGA), we estimated the activation states of these transcriptomic processes in individual tumors. Finally, we applied individualized causal network learning to discover an ICN within each tumor. Our results show that cellular states of cells in TMEs are coordinated through ICNs that enable multi-way communications among epithelial, fibroblast, endothelial, and immune cells. Further analyses of individual ICNs revealed structural patterns that were shared across subsets of tumors, leading to the discovery of 4 different subtypes of networks that underlie disparate TMEs of HNSCC. Patients with distinct TMEs exhibited significantly different clinical outcomes. Our results show that the capability of estimating individual ICNs reveals heterogeneity of ICNs and sheds light on the importance of intercellular communication in impacting disease development and progression.

cancer biology↗

bioRxiv2021

Myofibroblast transcriptome indicates SFRP2+ fibroblast progenitors in systemic sclerosis skin

Skin and lung fibrosis in systemic sclerosis (SSc) is driven by myofibroblasts, alpha-smooth muscle actin expressing cells that arise from a variety of cell types in murine fibrosis models. Utilizing single cell RNA-sequencing to examine the transcriptome changes, we show that SSc dermal myofibroblasts arise from an SFRP2/DPP4-expressing progenitor fibroblast population that globally upregulates expression of transcriptome markers, such as PRSS23 and THBS1. Only a fraction of SSc fibroblasts differentiate into myofibroblasts, as shown by expression of additional markers, SFRP4 and FNDC1. The myofibroblast transcriptome implicates upstream transcription factors that drive myofibroblast differentiation.

cell biology↗

bioRxiv2021

Adipocyte lipolysis abrogates skin fibrosis in a Wnt/DPP4-dependent manner

Tissue fibrosis in many organs results from altered and excessive extracellular matrix (ECM) protein deposition 1. Concomitant with ECM expansion, resident lipid-filled cells including mature adipocytes are lost in human and mouse fibrosis2-5, yet the mechanisms that drive mature adipocyte lipid loss and their contribution to tissue fibrosis are unknown. Here, we identify an early, fibro-protective role of mature adipocyte lipolysis driven by Wnt signaling during fibrosis onset. Using chemical and genetic mouse models of skin fibrosis, we show that fibrotic stimuli induce and maintain lipolysis in mature dermal adipocytes. Loss of the lipolytic rate-limiting enzyme adipocyte triglyceride lipase (ATGL)6,7 in murine dermal adipocytes exacerbates bleomycin-induced fibrosis development. Adipocyte lipolysis is stimulated in the early stages of Wnt signaling-induced skin fibrosis and by Wnt agonists in vitro. Furthermore, deletion or inhibition of the Wnt target gene, CD26/Dipeptidyl peptidase 4 (DPP4) prevented Wnt-induced lipolysis and skin fibrosis in mice. Notably, DPP4 expression correlates with skin fibrosis severity in human patients. Thus, we propose that adipocyte-derived fatty acids and the Wnt-DPP4 axis act as essential regulators of ECM homeostasis within tissues and provide a therapeutic avenue to manipulate fibrosis.

cell biology↗

bioRxiv2020

Divergent cancer etiologies drive distinct B cell signatures and tertiary lymphoid structures

Current immunotherapy paradigms aim to reinvigorate CD8+ T cells, but the contribution of humoral immunity to antitumor immunity remains understudied1,2. Head and neck squamous cell carcinoma (HNSCC) is caused by either human papillomavirus (HPV+) or environmental carcinogens (i.e. tobacco and alcohol; HPV-)3,4. Here, we demonstrate that HPV+ HNSCC patients have transcriptional signatures of germinal center (GC) tumor infiltrating B cells (TIL-Bs) and spatial organization of immune cells consistent with GC-like tertiary lymphoid structures (TLS), both of which correlate with favorable outcomes in HNSCC patients. Further, our single-cell RNAseq data also indicate that GC TIL-Bs are characterized by distinct waves of gene expression consistent with dark zone, light zone and a transitional state of GC B cells. High-dimensional spectral flow cytometry permitted in depth characterization of activated, memory and GC TIL-Bs. Further, single cell RNAseq analysis and subsequent protein validation identified a role for semaphorin 4a (Sema4a) in the differentiation of GC TIL-Bs and indicated that expression of Sema4a was enhanced on GC TIL-Bs and within GC-like TLS in the TME. Thus, in contrast to some reports on the detrimental role of TIL-Bs in human tumors, our findings suggest that TIL-Bs play an instrumental role in antitumor immunity5,6. Novel therapeutics to enhance TIL-B responses in HNSCC should be prioritized as a compliment to current T-cell mediated immunotherapies.

immunology↗

bioRxiv2018

BAMM-SC: A Bayesian mixture model for clustering droplet-based single cell transcriptomic data from population studies

The recently developed droplet-based single cell transcriptome sequencing (scRNA-seq) technology makes it feasible to perform a population-scale scRNA-seq study, in which the transcriptome is measured for tens of thousands of single cells from multiple individuals. Despite the advances of many clustering methods, there are few tailored methods for population-scale scRNA-seq studies. Here, we have developed a BAyesiany Mixture Model for Single Cell sequencing (BAMM-SC) method to cluster scRNA-seq data from multiple individuals simultaneously. Specifically, BAMM-SC takes raw data as input and can account for data heterogeneity and batch effect among multiple individuals in a unified Bayesian hierarchical model framework. Results from extensive simulations and application of BAMM-SC to in-house scRNA-seq datasets using blood, lung and skin cells from humans or mice demonstrated that BAMM-SC outperformed existing clustering methods with improved clustering accuracy and reduced impact from batch effects. BAMM-SC has been implemented in a user-friendly R package with a detailed tutorial available on www.pitt.edu/~Cwec47/singlecell.html.

bioinformatics↗