Biology subjects

Shah, P.

Publications and source records attributed to Shah, P..

At least 73 records · Page 4Linked to original sources

bioRxiv2021

Polyamines and eIF5A hypusination facilitate SREBP2 translation and cholesterol synthesis to enhance enterovirus attachment and infection

Metabolism is key to cellular processes that underlie the ability of a virus to productively infect. Polyamines are small metabolites vital for many host cell processes including proliferation, transcription, and translation. Polyamine depletion also inhibits virus infection via diverse mechanisms, including inhibiting polymerase activity and viral translation. We showed that Coxsackievirus B3 (CVB3) attachment requires polyamines; however, the mechanism was unknown. Here, we report polyamines involvement in translation, through a process called hypusination, promotes expression of cholesterol synthesis genes by supporting SREBP2 translation, the master transcriptional regulator of cholesterol synthesis genes. Measuring bulk transcription, we found polyamines support expression of cholesterol synthesis genes, regulated by SREBP2. Polyamine depletion inhibits CVB3 by depleting cellular cholesterol. Exogenous cholesterol rescues CVB3 attachment, and mutant CVB3 resistant to polyamine depletion exhibits resistance to cholesterol perturbation. This study provides a novel link between polyamine and cholesterol homeostasis, a mechanism through which polyamines impact CVB3 infection.

microbiology↗

bioRxiv2021

SARS-Cov-2 Spike binding to ACE2 is stronger and longer ranged with glycans

Highly detailed steered Molecular Dynamics simulations are performed on differently glycosylated receptor binding domains of the SARS-CoV-2 spike protein. The binding strength and the binding range increases with glycosylation. The interaction energy rises very quickly with pulling the proteins apart and only slowly drops at larger distances. We see a catch slip type behavior where interactions during pulling break and are taken over by new interactions forming. The dominant interaction mode are hydrogen bonds but Lennard-Jones and electrostatic interactions are relevant as well. Statement of SignificanceGlycosylation of the receptor binding domain of the Spike protein of SARS-CoV-2 as well as the ACE2 receptor leads to stronger and longer ranged binding interactions between the proteins. Particularly, at shorter distances the interactions are between residues of the proteins themselves whereas at larger distances these interactions are mediated by the glycans. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=164 SRC="FIGDIR/small/452507v2_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@7b9021org.highwire.dtl.DTLVardef@338aadorg.highwire.dtl.DTLVardef@1d5921eorg.highwire.dtl.DTLVardef@6c27e7_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2021

Low lamin A levels enhance confined cell migration and metastatic capacity in breast cancer

Aberrations in nuclear size and shape are commonly used to identify cancerous tissue. However, it remains unclear whether the disturbed nuclear structure directly contributes to the cancer pathology or is merely a consequence of other events occurring during tumorigenesis. Here, we show that highly invasive and proliferative breast cancer cells have lower expression of the nuclear envelope proteins lamin A/C, leading to increased nuclear deformability that permits enhanced cell migration through confined environments that mimic interstitial spaces encountered during metastasis. Importantly, increasing lamin A/C expression in highly invasive breast cancer cells altered expression of numerous other proteins implicated in metastatic progression. Further supporting an important role of lamins in breast cancer metastasis, analysis of lamin levels in human breast tumors revealed a significant association between lower lamin A levels and decreased disease-free survival. These findings suggest that downregulation of lamin A/C may influence both biochemical and physical properties of the cell to promote breast cancer metastatic progression.

cancer biology↗

bioRxiv2021

riboviz 2: A flexible and robust ribosome profiling data analysis and visualization workflow

MotivationRibosome profiling, or Ribo-seq, is the state of the art method for quantifying protein synthesis in living cells. Computational analysis of Ribo-seq data remains challenging due to the complexity of the procedure, as well as variations introduced for specific organisms or specialized analyses. Many bioinformatic pipelines have been developed, but these pipelines have key limitations in terms of functionality or usability. ResultsWe present riboviz 2, an updated riboviz package, for the comprehensive transcript-centric analysis and visualization of Ribo-seq data. riboviz 2 includes an analysis workflow built on the Nextflow workflow management system, combining freely available software with custom code. The package is extensively documented and provides example configuration files for organisms spanning the domains of life. riboviz 2 is distinguished by clear separation of concerns between annotation and analysis: prior to a run, the user chooses a transcriptome in FASTA format, paired with annotation for the CDS locations in GFF3 format. The user is empowered to choose the relevant transcriptome for their biological question, or to run alternative analyses that address distinct questions. riboviz 2 has been extensively tested on various library preparation strategies, including multiplexed samples. riboviz 2 is flexible and uses open, documented file formats, allowing users to integrate new analyses with the pipeline. Availabilityriboviz 2 is freely available at github.com/riboviz/riboviz. Supplementary information

bioinformatics↗

bioRxiv2021

The Hypoxia Response Pathway Promotes PEP Carboxykinase Expression And Gluconeogenesis

Actively dividing cells, including some cancers, rely on aerobic glycolysis rather than oxidative phosphorylation to generate energy, a phenomenon termed "the Warburg effect1." Constitutive activation of the Hypoxia Inducible Factor (HIF-1), a transcription factor known for mediating an adaptive response to oxygen deprivation (hypoxia), is a hallmark of the Warburg effect2. HIF-1 is thought to promote glycolysis and suppress oxidative phosphorylation. Here, we show instead that HIF-1 can promote gluconeogenesis. Using a multiomics approach, we determined the genomic, transcriptomic, and metabolomic landscapes regulated by constitutively active HIF-1 in C. elegans. We performed RNA-seq and ChIP-seq under aerobic conditions in mutants lacking EGL-9, a key negative regulator of HIF-1, and then integrated these approaches to identify over a hundred genes directly and functionally upregulated by HIF-1. We show that HIF-1 directly promotes the expression of PCK-1, a PEP carboxykinase that is a rate-limiting mediator of gluconeogenesis3. This activation of PCK-1 by HIF-1 promotes survival in response to both oxidative and hypoxic stress. Our work is the first to identify functional direct targets of HIF-1 in vivo, and it describes the first complete metabolome induced by constitutive HIF-1 activation in any organism.

genetics↗

bioRxiv2021

Thiophenesulfonamides are specific inhibitors of quorum sensing in pathogenic Vibrios

Vibrio bacteria are pathogens of fish, shellfish, coral, and humans due to contaminated seafood consumption. Vibrio virulence factors are controlled by the cell-to-cell communication called quorum sensing, thus this signaling system is a promising target for therapeutic design. We screened a compound library and identified nine compounds, including several 2-thiophenesulfonamides, that inhibit the master quorum sensing transcription factor LuxR in Vibrio campbellii but do not affect cell growth. We synthesized a panel of 50 thiophenesulfonamide compounds to examine the structure-activity relationship effects on quorum sensing in vivo. The most potent molecule identified, PTSP (3-phenyl-1-(thiophen-2-ylsulfonyl)-1H-pyrazole), specifically inhibits LuxR homologs in multiple strains of Vibrio vulnificus, Vibrio parahaemolyticus, and V. campbellii with sub-micromolar concentrations. PTSP efficacy is driven by amino acid conservation in the binding pocket, which is accurately predicted using in silico modeling of inhibitors. Our results underscore the potential for developing thiophenesulfonamides as specific quorum sensing-directed treatments for Vibrio infections.

microbiology↗

bioRxiv2021

Promoter-sequence determinants and structural basis of primer-dependent transcription initiation in Escherichia coli

Chemical modifications of RNA 5' ends enable "epitranscriptomic" regulation, influencing multiple aspects of RNA fate. In transcription initiation, a large inventory of substrates compete with nucleoside triphosphates (NTPs) for use as initiating entities, providing an ab initio mechanism for altering the RNA 5' end. In Escherichia coli cells, RNAs with a 5'-end hydroxyl are generated by use of dinucleotide RNAs as primers for transcription initiation, "primer-dependent initiation." Here we use massively systematic transcript end readout ("MASTER") to detect and quantify RNA 5' ends generated by primer-dependent initiation for ~410 (~1,000,000) promoter sequences in E. coli. The results show primer-dependent initiation in E. coli involves any of the 16 possible dinucleotide primers and depends on promoter sequences in, upstream, and downstream of the primer binding site. The results yield a consensus sequence for primer-dependent initiation, YTSS-2NTSS-1NTSSWTSS+1, where TSS is the transcription start site, NTSS-1NTSS is the primer binding site, Y is pyrimidine, and W is A or T. Biochemical and structure-determination studies show that the base pair (nontemplate-strand base:template-strand base) immediately upstream of the primer binding site (Y:RTSS-2, where R is purine) exerts its effect through the base on the DNA template strand (RTSS-2) through inter-chain base stacking with the RNA primer. Results from analysis of a large set of natural, chromosomally-encoded E. coli promoters support the conclusions from MASTER. Our findings provide a mechanistic and structural description of how TSS-region sequence hard-codes not only the TSS position, but also the potential for epitranscriptomic regulation through primer-dependent transcription initiation.

biochemistry↗

bioRxiv2021

The landscape of transcriptional and translational changes over 22 years of bacterial adaptation

Organisms can adapt to an environment by taking multiple mutational paths. This redundancy at the genetic level, where many mutations have similar phenotypic and fitness effects, can make untangling the molecular mechanisms of complex adaptations difficult. Here we use the E. coli long-term evolution experiment (LTEE) as a model to address this challenge. To bridge the gap between disparate genomic changes and parallel fitness gains, we characterize the landscape of transcriptional and translational changes across 11 replicate populations evolving in parallel for 50,000 generations. By quantifying absolute changes in mRNA abundances, we show that not only do all evolved lines have more mRNAs but that this increase in mRNA abundance scales with cell size. We also find that despite few shared mutations at the genetic level, clones from replicate populations in the LTEE are remarkably similar to each other in their gene expression patterns at both the transcriptional and translational levels. Furthermore, we show that the bulk of the expression changes are due to changes at the transcriptional level with very few translational changes. Finally, we show how mutations in transcriptional regulators lead to consistent and parallel changes in the expression levels of downstream genes, thereby linking genomic changes to parallel fitness gains in the LTEE. These results deepen our understanding of the molecular mechanisms underlying complex adaptations and provide insights into the repeatability of evolution.

evolutionary biology↗

bioRxiv2021

Glycogen Synthase Kinase 3 regulates the genesis of the rare displaced ganglion cell retinal subtype

Glycogen Synthase Kinase 3 (GSK) proteins (GSK3 and GSK3{beta}) are key mediators of signaling pathways, with crucial roles in coordinating fundamental biological processes during neural development. Here we show that the complete loss of GSK3 signaling in mouse retinal progenitors leads to microphthalmia with broad morphological defects. Both proliferation of retinal progenitors and neuronal differentiation are impaired and result in enhanced cell death. A single wild-type allele of either Gsk3 or Gsk3{beta} is able to rescue these phenotypes. In this genetic context, all cell types are present with a functional retina. However, we unexpectedly detect a large number of cells in the inner nuclear layer expressing retinal ganglion cell (RGC)-specific markers (called displaced RGCs, dRGCs) when at least one allele of Gsk3 is expressed. Excess dRGCs lead to increased number of axons projecting into the ipsilateral medial terminal nucleus, an area of the brain belonging to the non-image-forming visual circuit and poorly targeted by RGCs in wild-type retina. Transcriptome analysis and optomotor response assay suggest that at least a subset of dRGCs in Gsk3 mutant mice are direction-selective RGCs. Our study thus uncovers a unique role of GSK3 in controlling the genesis of dRGCs, a rare and poorly characterized retinal cell type.

neuroscience↗

bioRxiv2021

The gastrin-releasing peptide regulates stress-enhanced fear and dopamine signaling

Fear extinction is an adaptive behavioral process critical for organisms survival, but deficiency in extinction may lead to PTSD. While the amygdala and its neural circuits are critical for fear extinction, the molecular identity and organizational logic of cell types that lie at the core of these circuits remain unclear. Here we report that mice deficient for amygdala-enriched gastrin-releasing peptide gene (Grp-/-) exhibit enhanced neuronal activity in the basolateral amygdala (BLA) and stronger fear conditioning, as well as deficient extinction in stress-enhanced fear learning (SEFL). rAAV2-retro-based tracing combined with visualization of the GFP knocked in the Grp gene showed that BLA receives GRPergic or conditioned stimulus projections from the indirect auditory thalamus-to-auditory cortex pathway, ventral hippocampus and ventral tegmental area. Transcription of dopamine-related genes was decreased in BLA of Grp-/- mice following SEFL extinction recall, suggesting that the GRP may mediate fear extinction regulation by dopamine. Impact statementMice deficient for the amygdala-enriched gastrin-releasing peptide gene are susceptible to stress-enhanced fear, a behavioral protocol with relevance to PTSD, and show a decrease in dopamine-related gene transcription.

neuroscience↗

bioRxiv2020

Implantable photonic neural probes for light-sheet fluorescence brain imaging

SignificanceLight-sheet fluorescence microscopy is a powerful technique for high-speed volumetric functional imaging. However, in typical light-sheet microscopes, the illumination and collection optics impose significant constraints upon the imaging of non-transparent brain tissues. Here, we demonstrate that these constraints can be surmounted using a new class of implantable photonic neural probes. AimMass manufacturable, silicon-based light-sheet photonic neural probes can generate planar patterned illumination at arbitrary depths in brain tissues without any additional micro-optic components. ApproachWe develop implantable photonic neural probes that generate light sheets in tissue. The probes were fabricated in a photonics foundry on 200 mm diameter silicon wafers. The light sheets were characterized in fluorescein and in free space. The probe-enabled imaging approach was tested in fixed and in vitro mouse brain tissues. Imaging tests were also performed using fluorescent beads suspended in agarose. ResultsThe probes had 5 to 10 addressable sheets and average sheet thicknesses < 16 m for propagation distances up to 300 m in free space. Imaging areas were as large as {approx} 240 m x 490 m in brain tissue. Image contrast was enhanced relative to epifluorescence microscopy. ConclusionsThe neural probes can lead to new variants of light-sheet fluorescence microscopy for deep brain imaging and experiments in freely-moving animals.

bioengineering↗

bioRxiv2020

Epidemiology of Dengue viral infection in Nepal: 2006-2019

BackgroundDengue is one of the newest emerging diseases in Nepal with increasing burden and geographic spread over the last 14 years. The main objective of this study was to explore the spatio-temporal epidemiological patterns of Dengue since its first report (2006) till 2019 in Nepal. MethodsThis study is a retrospective analysis of dengue data available from the Epidemiological Disease Control Division (EDCD) of Government of Nepal. The data in this study cover the last 14 years (2006-2019) of reported dengue cases in Nepal. Epidemiological trend and spatio-temporal analyses were performed. Maps of reported case incidence were created using QGIS version 3.4. ResultsSince the first report of dengue in a foreigner in 2004, Nepal reported a total of 17,992 dengue cases in 68 districts of Nepal in 2019. The incidence was approximately five times higher in 2018 (Incidence Rate Ratio (IRR): 4.8; 95% CI: 1.5 - 15.3) and over 140 times higher in 2019 (IRR: 141.6; CI: 45.8 - 438.4). Population density was not a statistically significant predictor of case incidence. Mean elevation had a negative association with case incidence. A one standard deviation increase in elevation was associated with a 90% decrease in reported case incidence (IRR: 0.10; CI: 0.01 - 0.20). However, the association with mean elevation varied across the years. In comparison to 2016, incidence was greater at higher elevations in 2018 (IRR: 22.7; CI: 6.0 - 86.1) and 2019 (IRR: 9.6; CI: 2.6 - 36.1). ConclusionThere is a high risk of dengue outbreak in the Terai region with increasing spread towards the mid-mountains and beyond as seen over the last 14 years. Urgent measures are required to increase the availability of diagnostics and resources to mitigate future dengue epidemics. Findings from this study can inform the spatio-temporal distribution of dengue and can help in resource allocation and priority setting for future epidemic. Author summaryDengue in humans is caused by four different serotypes (DENV-1, DENV-2, DENV-3 & DENV-4). Globally it is the most pervasive vector borne diseases with increasing number of cases in recent years. Dengue is one of the youngest emerging diseases in Nepal with increasing cases and spread from the tropical lowland to the highland (hilly) regions. We conducted a spatio-temporal analysis of national data to consolidate the information using QGIS to measure the dengue incidence at district levels of Nepal. Spatio-temporal analysis exploring the incidence and distribution of dengue cases aids in identification of high-risk areas which can ultimately enable national dengue programme to mobilize and allocate resources for the control and treatment. This study shows, the persistent high risk of dengue outbreak in lowland Terai region with annual rise in the risk of spread towards the mid-mountains and beyond. Urgent measures are required to increase the diagnostics and resources to mitigate the epidemic burden of dengue in Terai and peripheral regions.

microbiology↗

bioRxiv2020

Structural basis for the neutralization of SARS-CoV-2 by an antibody from a convalescent patient

The COVID-19 pandemic has had unprecedented health and economic impact, but currently there are no approved therapies. We have isolated an antibody, EY6A, from a late-stage COVID-19 patient and show it neutralises SARS-CoV-2 and cross-reacts with SARS-CoV-1. EY6A Fab binds tightly (KD of 2 nM) the receptor binding domain (RBD) of the viral Spike glycoprotein and a 2.6[A] crystal structure of an RBD/EY6A Fab complex identifies the highly conserved epitope, away from the ACE2 receptor binding site. Residues of this epitope are key to stabilising the pre-fusion Spike. Cryo-EM analyses of the pre-fusion Spike incubated with EY6A Fab reveal a complex of the intact trimer with three Fabs bound and two further multimeric forms comprising destabilized Spike attached to Fab. EY6A binds what is probably a major neutralising epitope, making it a candidate therapeutic for COVID-19.

microbiology↗

bioRxiv2020

Nuclear deformation causes DNA damage by increasing replication stress

Cancer metastasis, i.e., the spreading of tumor cells from the primary tumor to distant organs, is responsible for the vast majority of cancer deaths. In the process, cancer cells migrate through narrow interstitial spaces substantially smaller in cross-section than the cell. During such confined migration, cancer cells experience extensive nuclear deformation, nuclear envelope rupture, and DNA damage. The molecular mechanisms responsible for the confined migration-induced DNA damage remain incompletely understood. While in some cell lines, DNA damage is closely associated with nuclear envelope rupture, we show that in others, mechanical deformation of the nucleus is sufficient to cause DNA damage, even in the absence of nuclear envelope rupture. This deformation-induced DNA damage, unlike nuclear envelope rupture-induced DNA damage, occurs primarily in S/G2 phase of the cell cycle and is associated with replication forks. Nuclear deformation, resulting from either confined migration or external cell compression, increases replication stress, possibly by increasing replication fork stalling, providing a molecular mechanism for the deformation-induced DNA damage. Thus, we have uncovered a new mechanism for mechanically induced DNA damage, linking mechanical deformation of the nucleus to DNA replication stress. This mechanically induced DNA damage could not only increase genomic instability in metastasizing cancer cells, but could also cause DNA damage in non-migrating cells and tissues that experience mechanical compression during development, thereby contributing to tumorigenesis and DNA damage response activation.

cell biology↗

bioRxiv2020

UBR-Box containing protein, UBR5, is over-expressed in human lung adenocarcinoma and is a potential therapeutic target

BackgroundN-end rule ubiquitination pathway is known to be disrupted in many diseases, including cancer. UBR5, an E3 ubiquitin ligase, is mutated and/or overexpressed in human lung cancer cells suggesting its pathological role in cancer. MethodsWe determined expression of UBR5 protein in multiple lung cancer cell lines and human patient samples. Using immunoprecipitation followed by mass spectrometry we determined the UBR5 interacting proteins. The impact of loss of UBR5 for lung adenocarcinoma cell lines was analyzed using cell viability, clonogenic assays and in vivo xenograft models in nude mice. Additional Western blot analysis was performed to assess the loss of UBR5 on downstream signaling. Statistical analysis was done by one-way ANOVA for in vitro studies and Wilcoxon paired t-test for tumor volumes in vivo. ResultsWe show variability of UBR5 expression levels in lung adenocarcinoma cell lines and in primary human patient samples. To gain better insight into the role that UBR5 may play in lung cancer progression we performed unbiased interactome analyses for UBR5. Data indicate that UBR5 has a wide range of interacting protein partners that are known to be involved in critical cellular processes such as DNA damage, proliferation and cell cycle regulation. We have demonstrated that shRNA-mediated loss of UBR5 decreases cell viability and clonogenic potential of lung adenocarcinoma cell lines. In addition, we found decreased levels of activated AKT signaling after the loss of UBR5 in lung adenocarcinoma cell lines using multiple means of UBR5 knockdown/knockout. Furthermore, we demonstrated that loss of UBR5 in lung adenocarcinoma cells results in significant reduction of tumor volume in nude mice. ConclusionsThese findings demonstrate that deregulation of the N-end rule ubiquitination pathway plays a crucial role in the etiology of some human cancers, and blocking this pathway via UBR5-specific inhibitors, may represent a unique therapeutic target for human cancers.

cancer biology↗

bioRxiv2020

Ubiquilin proteins regulate EGFR levels and activity in lung adenocarcinoma cells

Ubiquilin proteins (UBQLNs) are involved in diverse cellular processes like ERAD (endoplasmic reticulum associated degradation), autophagy, apoptosis and epithelial to mesenchymal transition. UBQLNs interact with a variety of substrates, including cell surface receptors, transcription factor regulators, proteasomal machinery proteins, and transmembrane proteins. Additionally, previous work from our lab shows that UBQLN1 interacts with IGFR family members (IGF1R, IGF2R, INSR) and this interaction regulates the activity and proteostasis of IGFR family members. Here, we examined regulation of UBQLN1 with Epidermal Growth Factor Receptor (EGFR) in lung adenocarcinoma cells. Loss of UBQLN1 occurs at high frequency in human lung cancer patient samples and we have shown that loss of UBQLN1 is capable altering processes involved in cell proliferation, migration, invasion and epithelial to mesenchymal transition in lung adenocarcinoma cell lines. Here, we present data that loss of UBQLN1 resulted in increased turnover of total EGFR, whilst increasing the relative amount of active EGFR in lung adenocarcinoma cells, especially in the presence of its ligand EGF. Furthermore, loss of UBQLN1 led to a more invasive cell phenotype as manifested by increased proliferation, migration and speed of movement of these lung adenocarcinoma cells. Taken together, UBQLN1 regulates expression and stability of IGFRs and EGFR, members of the receptor tyrosine kinase family of proteins in lung cancer cells.

cancer biology↗

bioRxiv2020

Ripasudil in a model of pigmentary glaucoma

PurposeTo investigate the effects of Ripasudil (K-115), a Rho-kinase inhibitor, in a porcine model of pigmentary glaucoma. MethodsHallmark features of trabecular meshwork (TM), the principle structure of the outflow system affected in this model, were analyzed. In vitro TM cells and ex vivo perfused eyes were subjected to pigment dispersion followed by K-115 treatment (PK115). PK115 was compared to sham-treated controls (C) and pigment (P). Cytoskeletal alterations were assessed by F-actin labeling. TM cell phagocytosis of fluorescent targets was evaluated by flow cytometry. Cell migration was studied with a wound-healing assay. Intraocular pressure was continuously monitored and compared to after the establishment of the pigmentary glaucoma model and after treatment with K-115. ResultsIn vitro, the percentage of cells with stress fibers increased in response to pigment but declined sharply after treatment with K-115 (P: 32.8 +/- 2.9%; PK115: 11.6 +/- 3.3%, P < 0.001). Phagocytosis first declined but recovered after K-115 (P: 25.7+/-2.1%, PK115: 33.4+/-0.8%, P <0.01). Migration recuperated at 12h with K-115 treatment (P: 19.1+/-4.6 cells/high-power field, PK115: 42.5+/-1.6 cells/high-power field, P <0.001). Ex vivo, eyes became hypertensive from pigment dispersion but were normotensive after treatment with K-115 (P: 20.3 +/- 1.2 mmHg, PK115: 8.9 +/- 1.7 mmHg; P< 0.005). ConclusionIn vitro, K-115 reduced TM stress fibers, restored phagocytosis, and restored migration of TM cells. Ex vivo, K-115 normalized intraocular pressure.

pharmacology and toxicology↗

bioRxiv2020

Comparative analysis of genome-wide DNA methylation identifies patterns that associate with conserved transcriptional programs in osteosarcoma.

Osteosarcoma is an aggressive tumor of the bone that primarily affects young adults and adolescents. Osteosarcoma is characterized by genomic chaos and heterogeneity. While inactivation of tumor suppressor p53 TP53 is nearly universal other high frequency mutations or structural variations have not been identified. Despite this genomic heterogeneity, key conserved transcriptional programs associated with survival have been identified across human, canine and induced murine osteosarcoma. The epigenomic landscape, including DNA methylation, plays a key role in establishing transcriptional programs in all cell types. The role of epigenetic dysregulation has been studied in a variety of cancers but has yet to be explored at scale in osteosarcoma. Here we examined genome-wide DNA methylation patterns in 24 human and 44 canine osteosarcoma samples identifying groups of highly correlated DNA methylation marks in human and canine osteosarcoma samples. We also link specific DNA methylation patterns to key transcriptional programs in both human and canine osteosarcoma. Building on previous work, we built a DNA methylation-based measure for the presence and abundance of various immune cell types in osteosarcoma. Finally, we determined that the underlying state of the tumor, and not changes in cell composition, were the main driver of differences in DNA methylation across the human and canine samples. SignificanceThis is the first large scale study of DNA methylation in osteosarcoma and lays the ground work for the exploration of DNA methylation programs that help establish conserved transcriptional programs in the context of different genomic landscapes.

cancer biology↗