Biology subjects

Grant, G. R.

Publications and source records attributed to Grant, G. R..

16 recordsLinked to original sources

bioRxiv2026

A Benchmark of Unsupervised Circadian Phase Inference Methods for Bulk Transcriptomic Data

Circadian phase can be inferred from gene expression when sample collection times are unavailable, but the available methods have not been systematically compared. We compared ten unsupervised phase inference methods and two clock-gene baselines on 25 datasets with reference times (16 real datasets from five studies and 9 simulations). The primary measure was the aligned circular mean absolute error (MAE), with a permutation test for significance, circular correlations and gene-level reconstruction of rhythmicity, acrophase and amplitude were secondary outcomes. CHIRAL had the lowest mean MAE (1.80 h, 95% confidence interval 1.21-2.97 h from resampling study families). The two clock-gene baselines (2.15 and 2.20 h) were next, followed by TEMPO, CYCLOPS and CYCLOPS 2.0 (2.49-2.80 h), and the confidence intervals of these six overlapped substantially. The other six methods had mean errors of 3.91-4.86 h, compared with 5.08-5.23 h for permuted phases. Accuracy depended on the dataset and on the strength of clock-gene rhythms, and, for several methods, on the random seed and the adjustment for batch or covariates. When the inferred phases were used to analyse gene rhythms in 22 datasets, the four leading methods identified rhythmic genes with a median ROC AUC of 0.64-0.70 and recovered gene acrophases with a median error of 0.18-0.64 h. In two mouse datasets without collection times, agreement with external reference acrophases differed between methods and tissues.

bioinformatics↗

bioRxiv2025

Role of IL10 signaling in the circadian control of host response to Influenza infection

We have previously demonstrated that the circadian clock regulates the host response to influenza A virus (IAV) infection, conferring a time-of-day-specific protection -infection at dawn resulted in a threefold increase in survival and reduced immunopathology compared to infection at dusk. While IL10 is well-known for its immunoregulatory function, its role in IAV remains unclear, with studies reporting both protective and detrimental effects. Given the diurnal rhythmicity of IL-10 receptor (Il10ra) expression in the lung, we investigated the contribution of IL-10 signaling to time-of-day-specific IAV protection. We found that blocking IL10 signaling abrogated the time of day protection, leading to increased immunopathology characterized by enhanced lymphocyte infiltration and global immune activation (transcriptomic analysis). Interestingly, while later, IL-10R blockade also eliminated the time-of-day difference in IAV outcomes, it improved the outcome of dusk-infected mice. Furthermore, natural killer (NK) cell depletion suppressed IL-10 levels in bronchoalveolar lavage, suggesting a role for NK cells in regulating IL-10 signaling. In conclusion, incorporating the circadian context has not only clarified the IL-10 role in IAV infection but also underscored the pivotal influence of circadian regulation on immune responses.

immunology↗

bioRxiv2025

Generating Correlated Data for Omics Simulation

Simulation of realistic omics data is a key input for benchmarking studies that help users obtain optimal computational pipelines. Omics data involves large numbers of measured features on each samples and these measures are generally correlated with each other. However, simulation too often ignores these correlations, perhaps due to the inconvenience and computational hurdles of doing so. To alleviate this, we describe in detail three approaches for quickly generating omics-scale data with correlated measures which mimic real data sets. These approaches all are based on a Gaussian copula approach with a covariance matrix that decomposes into a diagonal part and a low-rank part. We use these approaches to demonstrate the importance of including correlation in two benchmarking applications. First, we show that variance of results from the popular DESeq2 method increases when dependence is included. Second, we demonstrate that CYCLOPS, a method for inferring circadian time of collection from transcriptomics, improves in performance when given gene-gene dependencies in some circumstances. We provide an R package, dependentsimr, that has efficient implementations of these methods and can generate dependent data with arbitrary distributions, including discrete (binary, ordered categorical, Poisson, negative binomial), continuous (normal), or with an empirical distribution.

bioinformatics↗

bioRxiv2025

Sources of non-uniform coverage in short-read RNA-Seq data

The origin of several normal cellular functions and related abnormalities can be traced back to RNA splicing. As such, RNA splicing is currently the focus of a vast array of studies. To quantify the transcriptome, short-read RNA-Seq remains the standard assay. The primary technical artifact of RNA-Seq library prep, which severely interferes with analysis, is extreme non-uniformity in coverage across transcripts. This non-uniformity is present in both bulk and single-cell RNA-Seq and is observed even when the sample contains only full-length transcripts. This issue dramatically affects the accuracy of isoform-level quantification of multi-isoform genes. Understanding the sources of this non-uniformity is critical to developing improved protocols and analysis methods. Here, we explore eight potential sources of non-uniformity. We demonstrate that it cannot be explained by one factor alone. We performed targeted experiments to investigate the effect of fragment length, PCR ramp rate, and ribosomal depletion. We assessed existing data sets with varying sample quality, PCR cycle number, reverse transcriptase, and technical or biological replicates. We found evidence that interference of reverse transcription by secondary structure is unlikely to be the major contributing factor, that rRNA pull-down methods do not cause non-uniformity, that PCR ramp rate does not substantially impact non-uniformity, and that shorter fragments do not reduce non-uniformity. All these findings contradict prior publications or recommendations.

bioinformatics↗

bioRxiv2024

Disruption of the PGE2 synthesis / response pathway restrains atherogenesis in programmed cell death-1 (Pd-1) deficient hyperlipidemic mice

Immune checkpoint inhibitors (ICIs) that target programmed cell death 1 (PD-1) have revolutionized cancer treatment by enabling the restoration of suppressed T-cell cytotoxic responses. However, resistance to single-agent ICIs limits their clinical utility. Combinatorial strategies enhance their antitumor effects, but may also enhance the risk of immune related adverse effects of ICIs. Prostaglandin (PG) E2, formed by the sequential action of the cyclooxygenase (COX) and microsomal PGE synthase (mPGES-1) enzymes, acting via its E prostanoid (EP) receptors, EPr2 and EPr4, promotes lymphocyte exhaustion, revealing an additional target for ICIs. Thus, COX inhibitors and EPr4 antagonists are currently being combined with ICIs potentially to enhance antitumor efficacy in clinical trials. However, given the cardiovascular (CV) toxicity of COX inhibitors, such combinations may increase the risk particularly of CV AEs. Here, we compared the impact of distinct approaches to disruption of the PGE2 synthesis /response pathway - global or myeloid cell specific depletion of mPges-1 or global depletion of Epr4 - on the accelerated atherogenesis in Pd-1 deficient hyperlipidemic (Ldlr-/-) mice. All strategies restrained the atherogenesis. While depletion of mPGES-1 suppresses PGE2 biosynthesis, reflected by its major urinary metabolite, PGE2 biosynthesis was increased in mice lacking EPr4, consistent with enhanced expression of aortic Cox-1 and mPges-1. Deletions of mPges-1 and Epr4 differed in their effects on immune cell populations in atherosclerotic plaques; the former reduced neutrophil infiltration, while the latter restrained macrophages and increased the infiltration of T-cells. Consistent with these findings, chemotaxis by bone-marrow derived macrophages from Epr4-/- mice was impaired. Epr4 depletion also resulted in extramedullary lymphoid hematopoiesis and inhibition of lipoprotein lipase activity (LPL) with coincident spelenomegaly, leukocytosis and dyslipidemia. Targeting either mPGES-1 or EPr4 may restrain lymphocyte exhaustion while mitigating CV irAEs consequent to PD-1 blockade.

pharmacology and toxicology↗

bioRxiv2023

BEERS2: RNA-Seq simulation through high fidelity in silico modeling

Simulation of RNA-seq reads is critical in the assessment, comparison, benchmarking, and development of bioinformatics tools. Yet the field of RNA-seq simulators has progressed little in the last decade. To address this need we have developed BEERS2, which combines a flexible and highly configurable design with detailed simulation of the entire library preparation and sequencing pipeline. BEERS2 takes input transcripts (typically fully-length mRNA transcripts with polyA tails) from either customizable input or from CAMPAREE simulated RNA samples. It produces realistic reads of these transcripts as FASTQ, SAM, or BAM formats with the SAM or BAM formats containing the true alignment to the reference genome. It also produces true transcript-level quantification values. BEERS2 combines a flexible and highly configurable design with detailed simulation of the entire library preparation and sequencing pipeline and is designed to include the effects of polyA selection and RiboZero for ribosomal depletion, hexamer priming sequence biases, GC-content biases in PCR amplification, barcode read errors, and errors during PCR amplification. These characteristics combine to make BEERS2 the most complete simulation of RNA-seq to date. Finally, we demonstrate the use of BEERS2 by measuring the effect of several settings on the popular Salmon pseudoalignment algorithm.

bioinformatics↗

bioRxiv2022

Meta-analysis of diurnal transcriptomics reveals strong patterns of concordance and discordance in mouse liver

The accumulation of public transcriptomic timeseries data enables robust meta-analyses that were not possible until recently. To assess the consistency of biological rhythms across studies, 43 public mouse liver tissue timeseries totaling 805 RNA-seq samples were obtained and analyzed. Only the control groups of each study were included, in order to create comparable data. Technical factors in RNA-seq library preparation were the largest contributors to transcriptome-level differences, beyond biological or experiment-specific factors such as lighting conditions. Core clock genes were remarkably consistent in phase across all studies, while phase distributions of other periodic genes were generally less consistent. Overlap of genes identified as rhythmic across studies was generally low, up to around 50% between some of the highest sample count studies. Distributions of phases of significant genes were remarkably inconsistent across studies, but genes consistently identified as rhythmic clustered near ZT0 and ZT12 in acrophase. Data was integrated across studies in a JIVE analysis, which showed that the top two components of joint within-study variation are determined by time of day. A shape-invariant model with random effects was fit to the genes to identify the underlying shape of the rhythms, consistent across all studies. This revealed the extent of asymmetric and multimodal genes.

genomics↗

bioRxiv2022

The Genetic and Evolutionary Basis of Gene Expression Variation in East Africans

BackgroundMapping of quantitative trait loci (QTL) associated with molecular phenotypes is a powerful approach for identifying the genes and molecular mechanisms underlying human traits and diseases. How the genetic architecture of molecular traits varies across human populations, however, has been less explored. To better understand the genetics of gene regulation in East Africans, we perform expression and splicing QTL mapping in whole blood from a cohort of 162 diverse Africans from Ethiopia and Tanzania. We assess replication of these QTLs in cohorts of predominantly European ancestry and identify candidate genes under selection in human populations. ResultsWe find the gene regulatory architecture of African and non-African populations is broadly shared, though there is a considerable amount of variation at individual loci across populations. Comparing our analyses to an equivalently sized cohort of European Americans, we find that QTL mapping in Africans improves the detection of expression QTLs and fine mapping of causal variation. Integrating our QTL scans with signatures of selection, we find several genes related to immunity and metabolism that are highly differentiated between Africans and non-Africans, as well as a gene associated with pigmentation, TMEM216, with evidence of population-specific selection in Nilo-Saharan speaking pastoralists. ConclusionExtending QTL-mapping studies beyond groups of European ancestry, particularly to diverse indigenous populations, is vital for a complete understanding of the genetic architecture of human traits and can reveal novel functional variation underlying human traits and disease.

genomics↗

bioRxiv2021

Circadian regulation of lung repair and regeneration

Optimal lung repair and regeneration is essential for recovery from viral infections such as that induced by influenza A virus (IAV). We have previously demonstrated that lung inflammation induced by IAV is under circadian control. However, it is not known if the circadian clock exerts its influence on lung repair and regenerative processes independent of acute inflammation from IAV. Here, we demonstrate for the first time that lung organoids have a functional clock as they mature and that the absence of an intact circadian clock impairs regenerative capacity. Using several models of circadian disruption, we show that with the absence of an intact clock lung proliferation is disrupted. Further, we find that the circadian clock acts through direct control of the Wnt/{beta}-catenin pathway. We speculate, that adding the circadian dimension to the critical process of lung repair and regeneration will lead to novel therapies and improve outcomes. Finally, we use data from UK Biobank to demonstrate at the population level, the role of poor circadian rhythms in mediating negative outcomes following lung infection.

cell biology↗

bioRxiv2021

RNA splicing analysis using heterogeneous and large RNA-seq1datasets

The ubiquity of RNA-seq has led to many methods that use RNA-seq data to analyze variations in RNA splicing. However, available methods are not well suited for handling heterogeneous and large datasets. Such datasets scale to thousands of samples across dozens of experimental conditions, exhibit increased variability compared to biological replicates, and involve thousands of unannotated splice variants resulting in increased transcriptome complexity. We describe here a suite of algorithms and tools implemented in the MAJIQ v2 package to address challenges in detection, quantification, and visualization of splicing variations from such datasets. Using both large scale synthetic data and GTEx v8 as benchmark datasets, we demonstrate that the approaches in MAJIQ v2 outperform existing methods. We then apply MAJIQ v2 package to analyze differential splicing across 2,335 samples from 13 brain subregions, demonstrating its ability to offer new insights into brain subregion-specific splicing regulation.

bioinformatics↗

bioRxiv2020

MOCCASIN: A method for correcting for known and unknown confounders in RNA splicing analysis

While the effects of confounders on gene expression analysis have been extensively studied there is a lack of equivalent analysis and tools for RNA splicing analysis. Here we assess the effect of confounders in two large public RNA-Seq datasets (TARGET, ENCODE), develop a new method, MOCCASIN, to correct the effect of both known and unknown confounders on RNA splicing quantification, and demonstrate MOCCASINs effectiveness on both synthetic and real data.

genomics↗

bioRxiv2020

Myeloid cell mPGES-1 mediates inflammatory pain hypersensitivity in mice.

Nonsteroidal anti-inflammatory drugs (NSAIDs) relieve inflammatory pain by predominant suppression of cyclooxygenase-2 derived prostaglandin (PG) E2. Innate immune cells contribute to inflammatory pain hypersensitivity and may be an attractive target for novel non-addictive approaches to pain management. We studied the contribution of PGE2 produced by myeloid cell microsomal prostaglandin E synthase -1 (mPGES-1) to peripheral inflammation and hyperalgesia in mice. Selective deletion of mPGES-1 in myeloid cells by crossing LysM-Cre mice with mPGES-1flox/flox mice (Mac-mPGES-1-KO) resulted in significantly reduced mechanical and thermal hyperalgesia in complete Freunds adjuvant (CFA)-evoked hind paw inflammation, zymosan-induced peri-articular inflammation and collagen II antibody-induced arthritis models. Systemic COX-2 inhibition or myeloid cell specific COX-2 deletion (by crossing LysM-Cre with COX-2 flox/flox mice) recapitulated reduction of CFA-induced inflammation and hyperalgesia. In contrast, deletion of mPGES-1 in neurons and glial cells by crossing mPGES-1flox/flox mice with Nestin-Cre mice had no detectable effect on inflammatory pain hypersensitivity. While macrophage recruitment was unaltered, tissue concentrations of PGE2, IL-1{beta} and TNF were significantly reduced in Mac-mPGES-1-KO paw tissues following CFA induction. Our results demonstrate that myeloid cell mPGES-1 is the dominant source of PGE2 in inflammatory pain hypersensitivity. Targeting myeloid cell mPGES-1 may afford a novel approach to inflammatory pain therapy.

pharmacology and toxicology↗

bioRxiv2020

Sex dependent compensatory mechanisms to preserve blood pressure homeostasis in PGI2 receptor deficient mice

Inhibitors of microsomal prostaglandin E synthase-1 (mPges-1) are in the early phase of clinical development. Deletion of mPges-1 confers analgesia, restrains atherogenesis and fails to accelerate thrombogenesis, while suppressing prostaglandin (PG) E2, but increasing biosynthesis of prostacyclin (PGI2). In hyperlipidemic mice, this last effect represents the dominant mechanism by which mPges-1 deletion restrains thrombogenesis, while suppression of PGE2 accounts for its anti-atherogenic effect. However, the impact of mPges-1 depletion on blood pressure (BP) in this setting remains unknown. To address how differential effects on PGE2 and PGI2 might modulate salt-evoked BP responses in the absence of mPges-1, we generated mice lacking the I prostanoid (Ipr) receptor or mPges-1 on a hyperlipidemic background caused by deletion of the low density lipoprotein receptor (Ldlr KOs). Here, mPges-1 depletion significantly increased the BP response to salt loading in male Ldlr KO mice, whereas, despite the direct vasodilator properties of PGI2, Ipr deletion suppressed it. Furthermore, combined deletion of the Ipr abrogated the exaggerated BP response in male mPges-1 KO mice. Suppression of PGE2 biosynthesis was enough to explain the exaggerated BP response to salt loading by either mPges-1/Ldlr depletion or by an MPGES-1 inhibitor in mice expressing human mPGES-1. However, the lack of a hypertensive response to salt in Ipr-deficient mice was attributable to reactive activation of the atrial natriuretic peptide pathway. Interestingly, these unexpected BP phenotypes were not observed in female mice fed a high salt diet. This is attributable to the protective effect of estrogen in Ldlr KO mice and in Ipr /Ldlr DKOs. Thus, estrogen compensates for a deficiency in PGI2 to maintain BP homeostasis in response to high salt in hyperlipidemic female mice. In males, by contrast, augmented formation of ANP plays a similar compensatory role, restraining hypertension and oxidant stress in the setting of Ipr depletion. Hyperlipidemic males on a high salt diet might be at risk of a hypertensive response to mPGES-1 inhibitors.

pharmacology and toxicology↗

bioRxiv2018

Variability in the Analgesic Response to Ibuprofen Following Third-Molar Extraction is Associated with Differences in Activation of the Cyclooxygenase Pathway

The analgesic efficacy of non-steroidal anti-inflammatory drugs (NSAIDs) has long been recognized to be limited by substantial interindividual variability in pain relief, but the underlying mechanisms are not well understood. We performed pain phenotyping, functional neuroimaging, pharmacokinetic/pharmacodynamic assessments, inflammation biomarkers, and gene expression profiling in healthy subjects who underwent surgical extraction of bony impacted third molars, in order to characterize factors associated with heterogeneity in response to ibuprofen.\n\nSubjects were treated with rapid-acting ibuprofen (400 mg; N=19) or placebo (N=10) in a randomized, double-blind design. Compared to placebo, ibuprofen-treated subjects exhibited greater reduction in pain scores, alterations in regional cerebral blood flow in brain regions associated with pain processing, and inhibition of ex vivo cyclooxygenase activity and urinary prostaglandin metabolite excretion as indices of biochemical drug action (p<0.05). As expected, ibuprofen-treated subjects could be stratified into partial responders (N=9, required rescue medication within the dosing interval) and complete responders (N=10, no rescue medication). This was also reflected by differences in pain scores (p<0.01) as early as 30 minutes following drug administration (p<0.05). Variability in analgesic efficacy was not associated with demographic or clinical characteristics, ibuprofen pharmacokinetics, metabolizing enzyme genotype, or the degree of cyclooxygenase inhibition by ibuprofen. However, complete responders had higher concentrations of inflammatory biomarkers in urine and serum, than partial responders. Specifically, a stable urinary prostaglandin E2 metabolite, serum TNF and IL-8 were higher in patients who did not require rescue medication compared those who did (p < 0.05). RNAseq gene expression analysis in PBMCs collected after surgery and ibuprofen administration showed enrichment of inflammation related pathways among genes differentially expressed (q < 0.2) between complete and partial responders\n\nThese findings suggest that patients who receive substantial pain relief from ibuprofen have a more pronounced activation of the prostanoid biosynthetic pathway and regulation of the inflammatory pain phenotype differs from those patients who are insufficiently treated with ibuprofen alone and may require an opioid or other therapeutic intervention.

genomics↗

bioRxiv2018

Distinct vascular genomic response of proton and gamma radiation.

Purpose. The cardiovascular biology of proton radiotherapy is not well understood. We aimed to compare the genomic dose-response to proton and gamma radiation of the mouse aorta to assess whether their vascular effects may diverge.\n\nMaterials and methods. We performed comparative RNA sequencing of the aorta following (4 hrs) total-body proton and gamma irradiation (0.5 - 200 cGy whole body dose, 10 dose levels) of conscious mice. A trend analysis identified genes that showed a dose response.\n\nResults. While fewer genes were dose-responsive to proton than gamma radiation (29 vs. 194 genes; q-value [≤] 0.1), the magnitude of the effect was greater. Highly responsive genes were enriched for radiation response pathways (DNA damage, apoptosis, cellular stress and inflammation; p-value [≤] 0.01). Gamma, but not proton radiation induced additionally genes in vasculature specific pathways. Genes responsive to both radiation types showed almost perfectly superimposable dose-response relationships.\n\nConclusions. Despite the activation of canonical radiation response pathways by both radiation types, we detected marked differences in the genomic response of the murine aorta. Models of cardiovascular risk based on photon radiation may not accurately predict the risk associated with proton radiation.

systems biology↗

bioRxiv2017

Complete Transcriptome Profiling Of Normal And Age-Related Macular Degeneration Eye Tissues Reveals Changes In Regulation Of Non-Coding RNA And Extreme Disregulation Of Anti-Sense Transcription

Strand specific RNA sequencing of retina and RPE-Choroid-Scleara (RCS) in age-related macular degeneration (AMD) and matched normal controls reveals striking impact on anti-sense transcription and changes in the regulation of non-coding RNA that has not previously been reported. Hundreds of genes, which do not express anti-sense transcripts in normal retina and RCS, demonstrate extreme anti-sense expression in AMD. And conversely anti-sense transcription is completely abrogated in many genes which express a high level of anti-sense transcripts in normal retina and RCS. Several pathways are very highly enriched in the upregulated anti-sense transcripts - in particular the EIF2 signaling pathway. These results call for a deeper investigation into anti-sense and noncoding RNA regulation in AMD and their potential as therapeutic targets.

genomics↗