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Grenier, J. K.

Publications and source records attributed to Grenier, J. K..

15 recordsLinked to original sources
bioRxiv2026

From Prompt to Pipeline: A Comparative Evaluation of Large Language Model Coding Agents for Reproducible Bioinformatics Pipeline Construction

Agentic coding systems are increasingly presented as a way to reduce the engineering burden of scientific software development. Bioinformatics is a strong test case for this claim because useful pipelines must combine domain-specific analysis choices, command-line software, sample metadata, workflow orchestration, container or HPC execution, and interpretable quality-control reporting. We evaluated three agentic systems - Biomni, Claude Code, and Codex - on the same task: constructing a Nextflow DSL2 pipeline for paired-end CUT&Tag data that included read QC, trimming, alignment, filtering, duplicate removal, signal track generation, per-sample and group-level peak calling, control-aware group merging, annotation, FRiP calculation, deepTools visualizations, and final MultiQC reporting. Each system received the same detailed CRAFT-style prompt and was assessed against a hand-coded reference pipeline developed by the authors. All three systems produced pipeline implementations that appeared plausible at the level of documentation and file structure, but none fully satisfied the requested analysis. The most consequential failure was shared: the agent-generated pipelines performed some form of group-level merging but did not produce the requested merged-group reporting outputs. Sample-level MultiQC reports also disagreed with the reference report. Codex was closest to the reference for primary mapped-read counts, although its total-read accounting and report structure still differed. Claude Code produced the broadest final report, but its mapping summary mixed stages and therefore could not be treated as numerically correct. Biomni produced the strongest subjective documentation, but its read-count agreement with the reference report was poor and several failures required substantial Nextflow expertise to diagnose. These results suggest that current coding agents can accelerate scaffolding, documentation, and routine implementation, but they do not eliminate the need for expert review in bioinformatics workflow construction. For complex sequencing workflows, prompts must specify not only the biological intent, but also the exact stage semantics, acceptance tests, metadata contracts, expected report sections, resource propagation rules, and failure criteria needed to distinguish a plausible pipeline from a correct one.

bioinformatics↗
bioRxiv2026

BLink-seq delivers population-scale haplotypes without long reads: a scalable framework for non-model genomics

Information about segregating haplotypes and structural variation (SV) can be extremely rich for a variety of applications in population genomics but remains largely inaccessible for many non-model species. Of the available methods, linked-read sequencing is especially promising for its low cost and scalability, but its adoption remains limited. One existing linked-read method is Haplotagging, which barcodes sequencing reads to reconstruct long molecules that encode haplotype information, with the potential to generate phased whole-genome data and detect structural variants. In this study, we present BLink-seq, a novel Haplotagging method that is compatible with standard short-read next-generation sequencing platforms, is locally reproducible with low-cost reagents, and is scalable for high-throughput sample processing. We optimized library preparation parameters, explored their relationship to linked-read library metrics, and validated phasing performance and structural variant detection in two evolutionary extremes: an experimental Drosophila melanogaster cross of inbred lines carrying known inversions, and four Atlantic silverside (Menidia menidia) parent-offspring trios sourced from highly outbred, wild-caught populations. We then applied our protocol to a cohort of 376 silversides to demonstrate its scalability and potential for SV detection and genotype imputation. Using BLink-seq, we generated chromosome-scale phased blocks and identified known inversions in both validation datasets. We discovered previously uncharacterized structural complexity within a known adaptive inversion on silverside chromosome 11, demonstrating that linked-read data can refine our understanding of SV architecture beyond what short reads alone can resolve. Finally, we provide a user guide for researchers interested in using BLink-seq.

genomics↗
bioRxiv2026

Metagenomic and transcriptomic signatures of periodontitis in companion dogs

BackgroundPeriodontitis (Perio) is a progressive oral disease characterized by inflammation and degradation of the periodontal apparatus and is associated with local and systemic morbidity including loss of teeth, cardiovascular disease, and diabetes mellitus, among others. Perio is highly prevalent in domestic canines and exhibits certain parallels in pathogenesis and pathophysiology to Perio in humans, although standard treatments are less effective. In both species, a complex interplay between oral microbiota and host immune response is implicated in the etiology of Perio but is not fully understood. ResultsUsing shotgun metagenomics and RNA-seq on oral samples from companion dogs, we identify features of the oral microbiome and host transcriptional profile that are associated with Perio and its progression. We observe differences in microbiota composition between Perio and non-Perio animals that are largely consistent with what has been described in humans but also identify several species that are distinctly associated with canine Perio. We observe an abrupt shift in host gene expression related to immune response and tissue structure that is associated with disease severity, specifically the progression from mild periodontal disease (PD) to more severe Perio and the initiation of clinical attachment loss. The gingival plaque microbiota exhibits a parallel dynamic, with distinct compositional profiles in mild, moderate, and severe PD. We then examine several of the known mechanistic components of the keystone pathogen hypothesis of PD, identifying specific commonalities between canine and human pathologies, including the involvement of Porphyromonas species and related virulence factors. Additionally, we show infiltration of gingival tissue by Porphyromonas and Tannerella spp. via fluorescence microscopy. Finally, we assess correlations between host gene expression and microbial metabolic pathways which suggest additional potential virulence factors. ConclusionsThis work elucidates the metagenomic and transcriptomic signatures of Perio in companion dogs with the goals of informing veterinary medicine, evaluating the potential of canines as a model organism for the study of Perio, and clarifying the relationship between Perio development and progression, the oral microbiota, and the localized host response. Our findings provide insight into the etiopathogenesis of canine Perio and its relationship to human Perio and suggest novel targets of potential translational interest.

microbiology↗
bioRxiv2026

Neuroepithelial reprogramming and ERBB vulnerability in canine acanthomatous ameloblastoma

Canine acanthomatous ameloblastoma (CAA) is a locally invasive benign oral neoplasm that is difficult to distinguish from canine oral squamous cell carcinoma (COSCC) due to overlapping clinical, radiologic, and histologic features. Although both tumors exhibit MAPK pathway activation, their mutational landscapes are distinct. Furthermore, previous studies using bulk RNA sequencing (RNA-seq) have demonstrated pronounced differences in programs related to, among others, hypoxia, PI3K-AKT signaling, and cell proliferation. However, these bulk studies lacked the resolution to elucidate the cellular heterogeneity of CAA relative to COSCC. We therefore performed single-nucleus RNA-seq to define the cellular gene expression landscape of CAA, COSCC, and healthy gingiva. Across [~]205,000 nuclei, we identified major epithelial, immune, endothelial, and mesenchymal populations, as well as two epithelial subtypes uniquely enriched in CAA. The CAA-specific keratinocytes exhibited a neuronal-like expression program defined by synaptic regulators, KRAS-associated signaling pathways, and markedly elevated expression of PEG3, ERBB4, GABRB1, MAGI2, and CASK. These findings were validated by bulk RNA-seq, qPCR, and immunohistochemistry, which demonstrated strong nuclear localization of PEG3 exclusively in CAA epithelium. A kinase inhibitor screen independently identified ERBB4 as a candidate therapeutic vulnerability, and pharmacologic inhibition with neratinib was effective. Together, these findings reveal a previously unrecognized neuroepithelial cell state that defines CAA, distinguishes it from COSCC, and reveals unique diagnostic and therapeutic signaling dependencies. Given the molecular and histopathologic parallels between CAA and human ameloblastoma, these data further position CAA as a naturally occurring comparative model for studying ameloblastoma biology and therapeutic vulnerabilities. Significance StatementCanine acanthomatous ameloblastoma (CAA) is a common locally aggressive oral neoplasm often misdiagnosed as canine oral squamous cell carcinoma (COSCC) due to overlapping clinical, radiologic, and histologic features. Using single-nucleus RNA sequencing of CAA, COSCC, and healthy gingiva, we resolve the microenvironment of these tumors and detect two epithelial subpopulations unique to CAA. These CAA-specific keratinocytes exhibit a neuroepithelial-like signature characterized by synaptic regulators, KRAS-associated signaling, and elevated PEG3 and ERBB4 expression. Functional kinase inhibitor screening of patient derived microtumor slices independently converged on ERBB4 as an effective therapeutic target. Together, our study provides a molecular framework for distinguishing CAA from COSCC and establishes a comparative oncology model with translational relevance to human ameloblastoma and other ERBB-driven epithelial neoplasms.

cancer biology↗
bioRxiv2025

Genealogical analysis of replicate flower colour hybrid zones in Antirrhinum

A major goal of speciation research is identifying loci that underpin barriers to gene flow. Population genomics takes a bottom-up approach, scanning the genome for molecular signatures of processes that drive or maintain divergence. However, interpreting the genomic landscape of speciation is complicated, because genome scans conflate multiple processes, most of which are not informative about gene flow. However, studying replicated population contrasts, including multiple incidences of secondary contact, can strengthen inferences. In this paper, we use linked-read sequencing (haplotagging), FST scans, and genealogical methods to characterise the genomic landscape associated with replicate hybrid zone formation. We studied two flower colour varieties of the common snapdragon, Antirrhinum majus subspecies majus, that form secondary hybrid zones in multiple independent valleys in the Pyrenees. Consistent with past work, we found very low differentiation at one well-studied zone (Planoles). However, at a second zone (Avallenet), we found stronger differentiation and greater heterogeneity, which we argue is due to differences in the amount of introgression following secondary contact. Topology weighting of genealogical trees identified loci where haplotype diversity was associated with the two snapdragon varieties. Two of the strongest associations were at previously identified flower colour loci: Flavia, that affects yellow pigmentation, and Rosea/Eluta, two linked loci that affect magenta pigmentation. Preliminary analysis of coalescence times provides additional evidence for selective sweeps at these loci and barriers to gene flow. Our study highlights the impact of demographic history on the differentiation landscape, emphasizing the need to distinguish between historical divergence and recent introgression.

evolutionary biology↗
bioRxiv2025

Single-cell multiomics reveals the distinct properties of neonatal and adult recent thymic emigrants

Following thymic egress, CD8+ T cells must undergo a post-thymic maturation process to transition from a recent thymic emigrant (RTE) to a mature naive T cell. Since the neonatal T cell pool is comprised of significantly more RTEs, the prevailing notion is that neonatal CD8+ T cells behave differently than their adult counterparts simply because they have undergone less post-thymic maturation. To test this theory, we leveraged a fate mapping mouse model and paired single cell transcriptome and TCR sequencing to compare neonatal and adult CD8+ RTEs that have undergone the same amount of post-thymic maturation. Interestingly, we found that neonatal and adult CD8+ RTEs exhibit distinct phenotypes, gene expression profiles, TCR usage, and functions. These data suggest that neonatal CD8+ T cells are not simply immature adult CD8+ T cells and that age-related changes in CD8+ T cell functions in early life cannot be attributed solely to differences in the amount of post-thymic maturation.

immunology↗
bioRxiv2025

Argonaute proteins regulate the timing of the spermatogenic transcriptional program

Argonaute proteins are best known for their role in microRNA-mediated post-transcriptional gene silencing. Here, we show that AGO3 and AGO4, but not AGO2, localize to the sex chromatin of pachytene spermatocytes where they are required for transcriptional silencing of XY-linked genes, known as Meiotic Sex Chromosome Inactivation (MSCI). Using an Ago413-/- mouse, we show that AGO3 and AGO4 are key regulators of spermatogenesis, orchestrating expression of meiosis-related genes during prophase I while maintaining silencing of spermiogenesis genes. Premature overexpression of spermiogenesis genes during prophase I in Ago413-/- mice results in subfertility, altered sperm morphology and reduced fertilization capability. We also identify BRG1, a BAF complex subunit, as an AGO3 interactor. Loss of AGO3 and AGO4 results in increased BRG1 in spermatocytes, suggesting that AGO3 aids in removing BRG1 from the XY chromatin to achieve MSCI and demonstrating a meiotic role for AGO3 in transcriptional control through the chromatin remodeling machinery.

developmental biology↗
bioRxiv2024

Single-cell multi-omic analysis of fibrolamellar carcinoma reveals rewired cell-to-cell communication patterns and unique vulnerabilities

Fibrolamellar carcinoma (FLC) is a rare malignancy disproportionately affecting adolescents and young adults with no standard of care. FLC is characterized by thick stroma, which has long suggested an important role of the tumor microenvironment. Over the past decade, several studies have revealed aberrant markers and pathways in FLC. However, a significant drawback of these efforts is that they were conducted on bulk tumor samples. Consequently, identities and roles of distinct cell types within the tumor milieu, and the patterns of intercellular communication, have yet to be explored. In this study we unveil cell-type specific gene signatures, transcription factor networks, and super-enhancers in FLC using a multi-omics strategy that leverages both single-nucleus ATAC-seq and single-nucleus RNA-seq. We also infer completely rewired cell-to-cell communication patterns in FLC including signaling mediated by SPP1-CD44, MIF-ACKR3, GDF15-TGFBR2, and FGF7-FGFR. Finally, we validate findings with loss-of-function studies in several models including patient tissue slices, identifying vulnerabilities that merit further investigation as candidate therapeutic targets in FLC.

cancer biology↗
bioRxiv2024

Deciphering gene regulatory programs underlying functionally divergent naive T cell subsets

Naive CD8+ T cells are a heterogeneous population, with different subsets possessing distinct functions and kinetics upon activation. However, the gene regulatory circuits differentiating these naive subsets are not well studied. In this work, we analyzed a large collection of public and newly generated RNA-seq and ATAC-seq profiles of different subsets of naive CD8+ T cells, revealing significant differences in the gene regulatory landscapes between subsets. We leveraged these data by employing a network inference algorithm, Inferelator, to identify the transcriptional regulatory circuits active in each subset. The predicted transcriptional network of the naive CD8+ T cell pool was validated by multiple orthogonal approaches, including CUT&Tag and Micro-C. Interestingly, our network analysis revealed a novel role for Eomes in promoting effector cell differentiation in specific cell subsets. Moreover, we uncovered multiple novel regulators across a variety of subsets and discovered several modules of genes that were co-regulated by shared sets of transcription factors in distinct subsets. Collectively, our data defines the gene regulatory programs differentiating naive CD8+ T cells and facilitates the identification of novel transcription factors that may alter the propensity of naive CD8+ T cells to become effector or memory cells after infection.

systems biology↗
bioRxiv2023

Detection and characterization of novel luchacoviruses, genus Alphacoronavirus, in meso-carnivores in the northeastern United States

Small to mid-sized carnivores, or meso-carnivores, comprise a group of diverse mammals, many of which can adapt to anthropogenically disturbed environments. Wild meso-carnivores living in urban areas may get exposed to or spread pathogens to other species, including stray/feral domestic animals. Several coronaviruses (CoVs) have been detected in domesticated and farmed meso-carnivores, but knowledge of CoVs circulating in free-ranging wild meso-carnivores remains limited. In this study, we analyzed 321 samples collected between 2016 and 2022 from 9 species of free-ranging wild meso-carnivores and stray/feral domestic cats in the northeastern United States. Using a pan-CoV PCR, we screened tissues, feces, and saliva, nasal, and rectal swabs. We detected CoV RNA in fecal and saliva samples of animals in four species: fisher (Pekania pennanti), bobcat (Lynx rufus), red fox (Vulpes vulpes), and domestic cat (Felis catus). Next-generation sequencing revealed that all these viruses belonged to the Luchacovirus subgenus (Alphacoronavirus genus), previously reported only in rodents and lagomorphs (i.e., rabbits). Genetic comparison of the 3-end of the genome ([~]12,000bp) revealed that although the viruses detected group with, and have a genetic organization similar to other luchacoviruses, they are genetically distinct from those from rodents and lagomorphs. Genetic characterization of the spike protein revealed that the meso-carnivore luchacoviruses do not have an S1/S2 cleavage motif but do have highly variable structural loops containing cleavage motifs similar to those identified in certain pathogenic CoVs. This study highlights the importance of characterizing the spike protein of CoVs in wild species for further targeted epidemiologic monitoring. ImportanceSeveral coronaviruses (CoVs) have been detected in domesticated, farmed, and wild meso-carnivores, causing a wide range of diseases, and infecting diverse species, highlighting their important but understudied role in the epidemiology of these viruses. Assessing the viral diversity hosted in wildlife species is essential to understand their significance in the cross-species transmission of CoVs. Our focus here was on CoV discovery in meso-carnivores in the Northeast USA as a potential "hotspot" area with high density of humans and urban wildlife. This study identifies novel alphacoronaviruses circulating in multiple free-ranging wild and domestic species in this area and explores their potential epidemiological importance based on regions of the Spike gene that are relevant for virus-host interactions.

microbiology↗
bioRxiv2022

Single-cell transcriptomics of the immune system in ME/CFS at baseline and following symptom provocation

ME/CFS is a serious and poorly understood disease. To understand immune dysregulation in ME/CFS, we used single-cell RNA-seq (scRNA-seq) to examine immune cells in cohorts of patients and controls. Post-exertional malaise (PEM), an exacerbation of symptoms following strenuous exercise, is a characteristic symptom of ME/CFS. Thus, to detect changes coincident with PEM, we also performed scRNA-seq on the same cohorts following exercise. At baseline, ME/CFS patients displayed dysregulation of classical monocytes suggestive of inappropriate differentiation and migration to tissue. We were able to identify both diseased and more normal monocytes within patients, and the fraction of diseased cells correlated with metrics of disease severity. Comparing the transcriptome at baseline and post-exercise challenge, we discovered patterns indicative of improper platelet activation in patients, with minimal changes elsewhere in the immune system. Taken together, these data identify immunological defects present at baseline in patients and an additional layer of dysregulation following exercise. HighlightsME/CFS is a debilitating disease with unknown causes. Here, we provide, for the first time, an extensive single cell resolution dataset detailing the gene expression programs of circulating immune cells of ME/CFS cases at baseline and after symptom provocation. We were able to detect robust dysregulation in certain immune cells from patients, with dysregulation of classical monocytes manifesting the strongest signal. Indeed, the fraction of aberrant monocytes in ME/CFS patients correlated with the degree of disease severity. Surprisingly, platelet transcriptomes were also altered in ME/CFS, and they were the only component of the immune system that showed large-scale changes following symptom provocation.

immunology↗
bioRxiv2022

HIV-1 latent infection triggers broader epigenomic and transcriptional changes in protein-coding and long non-coding RNAs than active infection of SupT1 cells

Latent HIV-1 infection poses a major challenge in complete viral remission and cure. HIV-1 latency is a multi-dimensional, dynamic process and many aspects of how the viral latency is established and maintained still remains incompletely characterized. Here, we have investigated the host chromatin organization and transcriptomic changes in active- and latently-infected SupT1 cells. We employed an in vitro model of HIV-1 latency in SupT1 cells using a dual-reporter virus, HIVGKO, which enables high purity sorting and characterization of active- and latently-infected cells. We found a significant divergence in chromatin organization and gene expression pattern between active and latent infection compared to uninfected cells. Latent infection results in a repressive reorganization of the host chromatin, while active infection leads to an overall increase in chromatin accessibility. A stronger correlation was also observed between chromatin accessibility and gene expression in latent infection, which was manifested in a greater alteration of the cellular transcriptome in latent than active infection, for both protein-coding and long-non-coding RNAs (lncRNAs). We identified a number of novel lncRNAs associated with either active and latent infection. A reversal in expression pattern of latency-associated lncRNAs following PMA-induced reactivation indicated their infection-state-specific expression and potential roles in HIV-1 latency. Taken together, this integrated, comparative study revealed that latent HIV-1 infection requires a substantially greater alteration in cellular epigenome and transcriptome. Understanding of the distinct cellular states conducive to active and latent infection may support devising strategies for specific modulation of host cellular functions as a curative intervention for HIV-1.

microbiology↗
bioRxiv2022

Disrupted development and altered hormone signaling in male Padi2/Padi4 double knockout mice

BackgroundPeptidylarginine deiminase enzymes (PADs) convert arginine residues to citrulline in a process called citrullination or deimination. Recently, two PADs, PAD2 and PAD4, have been linked to hormone signaling in vitro and the goal of this study was to test for links between PAD2/PAD4 and hormone signaling in vivo. MethodsPreliminary analysis of Padi2 and Padi4 single knockout (SKO) mice did not find any overt reproductive defects and we predicted that this was likely due to genetic compensation. To test this hypothesis, we created a Padi2/Padi4 double knockout (DKO) mouse model and tested these mice along with wild-type FVB/NJ (WT) and both strains of SKO mice for a range of reproductive defects. ResultsControlled breeding trials found that male DKO mice appeared to take longer to have their first litter than WT controls. This tendency was maintained when these mice were mated to either DKO and WT females. Additionally, unsexed 2-day old DKO pups and male DKO weanlings both weighed significantly less than their WT counterparts, took significantly longer than WT males to reach puberty, and had consistently lower serum testosterone levels. Furthermore, 90-day old adult DKO males had smaller testes than WT males with increased rates of germ cell apoptosis. ConclusionsThe Padi2/Padi4 DKO mouse model provides a new tool for investigating PAD function and outcomes from our studies provide the first in vivo evidence linking PADs with hormone signaling.

molecular biology↗
bioRxiv2018

Dynamic transcriptome profiles within spermatogonial and spermatocyte populations during postnatal testis maturation revealed by single-cell sequencing

Spermatogenesis is the process by which male gametes are formed from a self-renewing population of spermatogonial stem cells (SSCs) residing in the testis. SSCs represent less than 1% of the total testicular cell population, but must achieve a stable balance between self-renewal and differentiation. Once differentiation has occurred, the newly formed and highly proliferative spermatogonia must then enter the meiotic program in which DNA content is doubled, then halved twice to create haploid gametes. While much is known about the critical cellular processes that take place during the specialized cell division that is meiosis, much less is known about how the spermatocytes in the \"first-wave\" compare to those that contribute to long-term, \"steady-state\" spermatogenesis. Given the strictly-defined developmental process of spermatogenesis, this study was aimed at exploring the transcriptional profiles of developmental cell stages over the age of the animal. Using a combination of comprehensive germ cell sampling with high-resolution, single-cell-mRNA-sequencing, we have generated a reference dataset of germ cell gene expression. We show that discrete developmental stages possess significant differences in the transcriptional profiles from neonates compared to juveniles and adults. Importantly, these gene expression dynamics are also reflected at the protein level in their respective cell types. We also show differential utilization of many biological pathways with age in both spermatogonia and spermatocytes, demonstrating significantly different underlying gene regulatory programs in these cell types over the course of testis development and spermatogenic waves. This dataset represents the first unbiased sampling of spermatogonia and spermatocytes in the developing testis over developmental age, at high-resolution, single-cell depth. Not only does this analysis reveal previously unknown transcriptional dynamics of a highly transitional cell population, it has also begun to reveal critical differences in biological pathway utilization in developing spermatogonia and spermatocytes, including response to DNA damage and double-strand breaks.\n\nAuthor SummarySpermatogenesis is the process by which male gametes - mature spermatozoa - are produced in the testis. This process requires exquisite control over many developmental transitions, including the self-renewal of the germline stem cell population, commitment to meiosis, and ultimately, spermiogenesis. While much is known about molecular mechanisms regulating single transitions at single time points in the mouse, much less is understood about how the spermatogenic progenitor cells, spermatogonia, or the meiotic cells, spermatocytes, of the testis change over developmental age.\n\nOur single-cell-mRNA-sequencing analysis is the first to profile both spermatogonia and spermatocytes from neonatal mice through adulthood, revealing novel gene expression dynamics and differential utilization of biological pathways. These discoveries help us to understand how the spermatogenic progenitors of this population modulate their activity to adapt to a changing testicular environment. Furthermore, they also begin to explain previously-observed differences - and deficiencies - in spermatocytes that are derived from the first \"wave\" of spermatogenesis. Overall, this dataset is the first of its kind to comprehensively profile gene expression dynamics in male germ cell populations over time, enriching our understanding of the complex and highly-orchestrated process of spermatogenesis.

genomics↗