Biology subjects

Allen, M. A.

Publications and source records attributed to Allen, M. A..

At least 19 recordsLinked to original sources

bioRxiv2026

Common cis-eQTLs do not explain chromosome 21 expression deviations from the ploidy expectation in Down syndrome

Down syndrome (DS) results from trisomy 21 (T21), the triplication of human chromosome 21 (HSA21). On average, genes on HSA21 are overexpressed at the expected 1.5-fold increase. A small subset of HSA21 genes is expressed closer to euploid levels. Could local genetic regulatory variation explain deviations from the expected dosage? Using the largest DS-specific transcriptomic dataset to date from the Human Trisome Project (HTP) and GTEx expression quantitative trait loci (eQTLs), we tested whether local genetic regulatory variation contributes to gene-specific buffering or amplification of HSA21 dosage effects in DS. We compared the bulk RNA-seq profiles of protein-coding genes, lncRNAs, and pseudogenes from whole-blood samples of 274 participants with T21 and 80 euploid (D21) participants. We filtered out genes with low read counts or high genomic repeat content and applied ploidy correction to the differential expression analysis. We found 13 genes that differed significantly from the expected fold change. We cross-referenced GTEx whole-blood eQTLs (cis window 1 {+/-} Mb) with the 13 deviating genes. A common cis-eQTL was detected for 8 of the 10 testable genes. However, 59 of 99 HSA21 genes that had the expected fold change had significant cis-eQTLs. Additionally, T21 participants carry the eQTL alleles at similar frequencies to GTEx donors. Therefore, cis-eQTLs explain differences between people with T21, not the difference between T21 and D21. Overall, common cis-regulatory variation acts on HSA21 genes in T21 as it does in euploid blood, but it does not explain which genes deviate from the ploidy expectation.

systems biology↗

bioRxiv2026

Attenuated estrogen signaling disrupts placentation and drives trophoblast defects in Down syndrome

Proper establishment of the primitive placenta and subsequent tissue homeostasis in the mature placenta are critical for successful pregnancy in humans. Placental insufficiency is associated with adverse pregnancy outcomes, including fetal growth restriction, preeclampsia, and pregnancy loss. Moreover, emerging evidence suggests that placental defects are associated with long-term health challenges that manifest well into adulthood; yet the etiologies of such diseases are largely unknown. Defining the mechanistic basis for placental deficiencies, therefore, has important implications for improving both reproductive health and the lifelong well-being of affected children. Down syndrome is characterized by placental defects of unknown mechanistic origin, and notably, individuals with Down syndrome are at increased risk of developing diseases commonly associated with placental insufficiency later in life. Using induced pluripotent stem cells from Down syndrome patients, we found that stem cell-based embryo models (i.e., blastoids) and directed differentiation systems recapitulate trophoblast cell fate defects observed in placentas affected by Down syndrome. Furthermore, we demonstrate that attenuated estrogen signaling contributes to placental syncytialization defects and identify NRIP1, a transcriptional corepressor of estrogen receptor that is located on chromosome 21, as a key driver of trophoblast cell fate defects. Increased gene dosage of NRIP1 in an otherwise diploid cell line phenocopies cell fate defects observed in trophoblasts affected by Down syndrome. Our study suggests that estrogen signaling is a crucial regulator of trophoblast development and may serve as a potential target for therapeutic intervention. Highlights and eTOC blurbO_LIEstrogen signaling mediates syncytiotrophoblast fusion C_LIO_LIHuman iPS cells provide a tractable model for trophoblast defects in Down syndrome C_LIO_LITrophoblast differentiation and estrogen signaling are disrupted in Down syndrome C_LIO_LIIncreased NRIP1 expression is sufficient to induce trophoblast defects C_LI Logsdon and colleagues apply patient-derived induced pluripotent stem cells to recapitulate placentation defects observed in Down syndrome. The authors demonstrate that attenuated estrogen signaling disrupts trophoblast differentiation and identify NRIP1, a gene found on chromosome 21 that dampens estrogen signaling, as a regulator of trophoblast maturation. NRIP1 and estrogen signaling may represent important therapeutic targets for infertility and Down syndrome.

developmental biology↗

bioRxiv2025

Base Composition Influences the Position andPrecision of RNA Polymerase II Disassociation inBasal and Perturbed Conditions.

RNA Polymerase II (Pol II) transcribes all protein-coding and many non-protein coding genes in the genome. Pol II transcription termination is crucial for mRNA maturation and, when disrupted, can lead to altered mRNA processing and mRNA export. Termination involves two intertwined processes: pre-mRNA cleavage and Pol II release from the DNA (disassociation). Despite its importance, the exact mechanisms underlying Pol II disassociation from the DNA remain poorly understood. Moreover, under certain cellular stress conditions, there is a partial failure of cleavage, leading to a shift of the position of disassociation further downstream, a phenomenon known as run-on transcription. We performed the first-ever systematic analysis of Pol II termination across cell types and species and provide novel insights into the mechanism of disassociation. Using a probabilistic mixture model to quantify Poll II dynamics across an entire gene body from nascent RNA sequencing data, we discovered that genes have two types of conserved regions near the disassociation site: one characterized by a T-rich region upstream of disassociation, and another characterized by a GC-rich region surrounding disassociation. Strikingly, the GC-rich disassociation regions have more accessible chromatin and higher levels of phospho-threonine 4 on the CTD of Pol II. Additionally, we find that upstream T-rich genes are preferentially affected by perturbations that alter disassociation, including heat-shock, viral infection, kinase inhibition, and arsenic treatment. Thus, our work has determined there are two types of Pol II disassociation regions, which are differentially affected by perturbation of cellular homeostasis.

genomics↗

bioRxiv2025

Improving confidence of differential transcription calls in enhancers

MotivationMost disease-associated genetic variants reside within transcribed regulatory elements (tREs). Patterns of differential transcription at tREs can be leveraged to identify upstream regulators and link enhancers to their target genes. But the low transcription levels and high variability in tREs makes identifying high confidence differentially transcribed elements challenging. ResultsWe present Mu Counts and TFEA-LE, two algorithms for robust identification of differentially transcribed tREs. The first step in accurate identification of differentially transcribed tREs is to obtain accurate RNA lengths and therefore counts over these regions. To this end we developed a method of accurate length inference (LIET-EMG) as wll as a rapid method for counting reads over tREs (Mu Counts). Armed with newly identified and quantified tREs, TFEA-LE then integrates motif information to simultaneously identify responsive tREs and their likely upstream regulators. We show improved precision and recall over general-purpose tools (e.g. DESeq2) in detecting p53-responsive tREs. We then clarify TF-specific responses within multi-TF perturbations in lung cells. Finally we show that the TFEA-LE approach improves TF activity inference, including in complex perturbations where many TFs respond. TFEALE is especially effective in technically challenging datasets, whether due to highly specific or broad responses, outliers, or high GC content. Ultimately, these 1 methods advance the systematic characterization of individual tREs, enabling their integration with regulators, target genes, and disease-associated variants for translational research. Availability and ImplementationTFEA-LE: https://github.com/Dowell-Lab/TFEA/tree/Lead_edge. Nextflow pipeline to run Mu Counts: https://github.com/Dowell-Lab/Bidir_Counting_Analysis. LIET (including modifications for tREs): https://github.com/Dowell-Lab/LIET/tree/LIET_EMGtoo. Source code for this work: https://github.com/Dowell-Lab/Improving_tRE_Analysis_Paper Contactrobin.dowell@colorado.edu

bioinformatics↗

bioRxiv2025

Histone Deacetylase Inhibitor Largazole Deactivates A Subset of Superenchancers and Causes Mitotic Chromosome Mis-alignment by Suppressing SP1 and BRD4

Histone deacetylase inhibitors have been investigated as potential therapeutic agents for cancer and other diseases. HDIs are known to promote histone acetylation, resulting in an open chromatin conformation and generally increased gene expression. In previous work, we reported that a subset of genes, particularly those regulated by superenhancers, can be suppressed by the HDAC inhibitor largazole. To elucidate the molecular mechanisms underlying gene repression by largazole, we conducted transposase-accessible chromatin sequencing, ChIP-seq, and RNA-seq studies. Our findings revealed that while largazole treatment generally enhances chromatin accessibility, it selectively decreases the accessibility of a subset of superenhancer regions. These genomic regions, showing the most significant changes in the presence of largazole, were enriched with transcription factor binding motifs for SP1, BRD4, CTCF, and YY1. ChIP-seq analysis confirmed reduced binding of BRD4 and SP1 at their respective sites on chromatin, particularly at superenhancers regulating genes such as ID1, c-Myc and MCMs. Largazole exerts its effects by inhibiting DNA replication, RNA processing, and cell cycle progression, partially through the suppression of SP1 expression. Depletion of SP1 by shRNA mimics several key biological effects of largazole and increases cellular sensitivity to the drug. Specific to cell cycle regulation, we demonstrated that largazole disrupts G/M transition by interfering with chromosome alignment during metaphase, a phenotype also observed with SP1 depletion. Our results suggest that largazole exerts its growth-inhibitory effect by suppressing BRD4 and SP1 at super-enhancers, leading to cytostatic responses and mitotic dysfunction.

cancer biology↗

bioRxiv2024

LIET Model: Capturing the kinetics of RNA polymerase from loading to termination

Transcription by RNA polymerases is an exquisitely regulated step of the central dogma. Transcription is the primary determinant of cell-state, and most cellular perturbations impact transcription by altering polymerase activity. Thus, detecting changes in polymerase activity yields insight into most cellular processes. Nascent run-on sequencing provides a direct readout of polymerase activity, but no tools exist to model this activity at genes. We focus on RNA polymerase II--responsible for transcribing protein-coding genes. We present the first model to capture the complete process of gene transcription. For individual genes, this model parameterizes each distinct stage of transcription--Loading, Initiation, Elongation, and Termination, hence LIET--in a biologically interpretable Bayesian mixture, which is applied to nascent run-on data. Our improved modeling of Loading /Initiation demonstrates these are characteristically different between sense and antisense strands. Applying LIET to 24 human cell-types, our analysis indicates the position of dissociation (the last step of Termination) appears to be highly consistent, indicative of a highly regulated process. Furthermore, applying LIET to perturbation experiments, we demonstrate its ability to detect specific changes in pausing (5'end), strand-bias, and dissociation location (3'end)--opening the door to differential assessment of transcription at individual stages of individual genes.

bioinformatics↗

bioRxiv2024

Single nucleus RNA-sequencing reveals transcriptional synchrony across different relationships

As relationships mature, partners share common goals, improve their ability to work together, and experience coordinated emotions. However, the neural underpinnings responsible for this unique, pair-specific experience remain largely unexplored. Here, we used single nucleus RNA-sequencing to examine the transcriptional landscape of the nucleus accumbens (NAc) in socially monogamous prairie voles in peer or mating-based relationships. We show that, regardless of pairing type, prairie voles exhibit transcriptional synchrony with a partner. Further, we identify genes expressed in oligodendrocyte progenitor cells that are synchronized between partners, correlated with dyadic behavior, and sensitive to partner separation. Together, our data indicate that the pair-specific social environment profoundly shapes transcription in the NAc. This provides a potential biological mechanism by which shared social experience reinforces and strengthens relationships.

neuroscience↗

bioRxiv2024

A transcription factor (TF) inference method that broadly measures TF activity and identifies mechanistically distinct TF networks

TF profiler is a method of inferring transcription factor regulatory activity, i.e. when a TF is present and actively regulating transcription, directly directly from nascent sequencing assays such as PRO-seq and GRO-seq. Transcription factors orchestrate transcription and play a critical role in cellular maintenance, identity and response to external stimuli. While ChIP assays have measured DNA localization, they fall short of identifying when and where transcription factors are actively regulating transcription. Our method, on the other hand, uses RNA polymerase activity to infer TF activity across hundreds of data sets and transcription factors. Based on these classifications we identify three distinct classes of transcription factors: ubiquitous factors that play roles in cellular homeostasis, driving basal gene programs across tissues and cell types, tissue specific factors that act almost exclusively at enhancers and are themselves regulated at transcription, and stimulus responsive TFs which are regulated post-transcriptionally but act predominantly at enhancers. TF profiler is broadly applicable, providing regulatory insights on any PRO-seq sample for any transcription factor with a known binding motif.

genomics↗

bioRxiv2023

Atlas of nascent RNA transcripts reveals enhancer to gene linkages

Gene transcription is controlled and modulated by regulatory regions, including enhancers and promoters. These regions are abundant in unstable, non-coding bidirectional transcription. Using nascent RNA transcription data across hundreds of human samples, we identified over 800,000 regions containing bidirectional transcription. We then identify highly correlated transcription between bidirectional and gene regions. The identified correlated pairs, a bidirectional region and a gene, are enriched for disease associated SNPs and often supported by independent 3D data. We present these resources as an SQL database which serves as a resource for future studies into gene regulation, enhancer associated RNAs, and transcription factors.

genomics↗

bioRxiv2023

Cellular zinc status alters chromatin accessibility and binding of transcription factor TP53 to genomic sites

Zinc (Zn2+) is an essential metal required by approximately 2500 proteins. Nearly half of these proteins act on DNA, including > 850 human transcription factors, polymerases, DNA damage response factors, and proteins involved in chromatin architecture. How these proteins acquire their essential Zn2+ cofactor and whether they are sensitive to changes in the labile Zn2+ pool in cells remain open questions. Here, we examine how changes in the labile Zn2+ pool affect chromatin accessibility and transcription factor binding to DNA. We observed both increases and decreases in accessibility in different chromatin regions via ATAC-seq upon treating MCF10A cells with elevated Zn2+ or the Zn2+-specific chelator tris(2-pyridylmethyl)amine (TPA). Transcription factor enrichment analysis was used to correlate changes in chromatin accessibility with transcription factor motifs, revealing 477 transcription factor motifs that were differentially enriched upon Zn2+ perturbation. 186 of these transcription factor motifs were enriched in Zn2+ and depleted in TPA, and the majority correspond to Zn2+ finger transcription factors. We selected TP53 as a candidate to examine how changes in motif enrichment correlate with changes in transcription factor occupancy by ChIP-qPCR. Using publicly available ChIP-seq and nascent transcription datasets, we narrowed the 50,000+ ATAC-seq peaks to 2164 TP53 targets and subsequently selected 6 high-probability TP53 binding sites for testing. ChIP-qPCR revealed that for 5 of the 6 targets, TP53 binding correlates with the local accessibility determined by ATAC-seq. These results demonstrate that changes in labile zinc directly alter chromatin accessibility and transcription factor binding to DNA.

genomics↗

bioRxiv2023

Mediator kinase inhibition suppresses hyperactive interferon signaling in Down syndrome

Hyperactive interferon (IFN) signaling is a hallmark of Down syndrome (DS), a condition caused by trisomy 21 (T21); strategies that normalize IFN signaling could benefit this population. Mediator-associated kinases CDK8 and CDK19 drive inflammatory responses through incompletely understood mechanisms. Using sibling-matched cell lines with/without T21, we investigated Mediator kinase function in the context of hyperactive IFN in DS over a 75min - 24h timeframe. Activation of IFN-response genes was suppressed in cells treated with the CDK8/CDK19 inhibitor cortistatin A (CA), via rapid suppression of IFN-responsive transcription factor (TF) activity. We also discovered that CDK8/CDK19 affect splicing, a novel means by which Mediator kinases control gene expression. To further probe Mediator kinase function, we completed cytokine screens and metabolomics experiments. Cytokines are master regulators of inflammatory responses; by screening 105 different cytokine proteins, we show that Mediator kinases help drive IFN-dependent cytokine responses at least in part through transcriptional regulation of cytokine genes and receptors. Metabolomics revealed that Mediator kinase inhibition altered core metabolic pathways in cell type-specific ways, and broad up-regulation of anti-inflammatory lipid mediators occurred specifically in kinase-inhibited cells during hyperactive IFN{gamma} signaling. A subset of these lipids (e.g. oleamide, desmosterol) serve as ligands for nuclear receptors PPAR and LXR, and activation of these receptors occurred specifically during hyperactive IFN signaling in CA-treated cells, revealing mechanistic links between Mediator kinases, lipid metabolism, and nuclear receptor function. Collectively, our results establish CDK8/CDK19 as context-specific metabolic regulators, and reveal that these kinases control gene expression not only via TFs, but also through metabolic changes and splicing. Moreover, we establish that Mediator kinase inhibition antagonizes IFN signaling through transcriptional, metabolic, and cytokine responses, with implications for DS and other chronic inflammatory conditions.

biochemistry↗

bioRxiv2023

Transcription dosage compensation does not occur in Down syndrome

BackgroundTrisomy 21, also known as Down syndrome, describes the genetic condition of having an extra copy of chromosome 21. The increase in DNA copy number has led to the "DNA dosage hypothesis", which claims that the level of gene transcription is proportional to the genes DNA copy number. Yet many reports have suggested that a proportion of chromosome 21 genes are dosage compensated back towards typical expression levels (1.0x). In contrast, other reports suggest that dosage compensation is not a common mechanism of gene regulation in Trisomy 21, providing support to the DNA dosage hypothesis. ResultsIn our work, we use both simulated and real data to dissect the elements of differential expression analysis that can lead to the appearance of dosage compensation even when compensation is demonstrably absent. Using lymphoblastoid cell lines derived from a family of an individual with Down syndrome, we demonstrate that dosage compensation is nearly absent at both nascent transcription (GRO-seq) and steady-state RNA (RNA-seq) levels. ConclusionsTranscriptional dosage compensation does not occur in Down syndrome. Simulated data containing no dosage compensation can appear to have dosage compensation when analyzed via standard methods. Moreover, some chromosome 21 genes that appear to be dosage compensated are consistent with allele specific expression.

bioinformatics↗

bioRxiv2023

The Dietary Fermentable Fiber Inulin Alters the Intestinal Microbiome and Improves Chronic Kidney Disease Mineral-Bone Disorder in a Rat Model of CKD

BackgroundDietary fiber is important for a healthy diet, but intake is low in CKD patients and the impact this has on the manifestations of CKD-Mineral Bone Disorder (MBD) is unknown. MethodsThe Cy/+ rat with progressive CKD was fed a casein-based diet of 0.7% phosphate with 10% inulin (fermentable fiber) or cellulose (non-fermentable fiber) from 22 weeks to either 30 or 32 weeks of age (~30 and ~15 % of normal kidney function). We assessed CKD-MBD, cecal microbiota, and serum gut-derived uremic toxins. Two-way ANOVA was used to evaluate the effect of age and inulin diet, and their interaction. ResultsIn CKD animals, dietary inulin led to changes in microbiota alpha and beta diversity at 30 and 32 weeks, with higher relative abundance of several taxa, including Bifidobacterium and Bacteroides, and lower Lactobacillus. Inulin reduced serum levels of gut-derived uremic toxins, phosphate, and parathyroid hormone, but not fibroblast growth factor-23. Dietary inulin decreased aorta and cardiac calcification and reduced left ventricular mass index and cardiac fibrosis. Bone turnover and cortical bone parameters were improved with inulin; however, bone mechanical properties were not altered. ConclusionsThe addition of the fermentable fiber inulin to the diet of CKD rats led to changes in the gut microbiota composition, lowered gut-derived uremic toxins, and improved most parameters of CKD-MBD. Future studies should assess this fiber as an additive therapy to other pharmacologic and diet interventions in CKD. Significance StatementDietary fiber has well established beneficial health effects. However, the impact of fermentable dietary fiber on the intestinal microbiome and CKD-MBD is poorly understood. We used an animal model of progressive CKD and demonstrated that the addition of 10% of the fermentable fiber inulin to the diet altered the intestinal microbiota and lowered circulating gut-derived uremic toxins, phosphorus, and parathyroid hormone. These changes were associated with improved cortical bone parameters, lower vascular calcification, and reduced cardiac hypertrophy, fibrosis and calcification. Taken together, dietary fermentable fiber may be a novel additive intervention to traditional therapies of CKD-MBD.

cell biology↗

bioRxiv2023

Characterizing Primary transcriptional responses to short term heat shock in paired fraternal lymphoblastoid lines with and without Down syndrome.

Heat shock stress induces genome wide changes in transcription regulation, activating a coordinated cellular response to enable survival. Using publicly available transcriptomic and proteomic data sets comparing individuals with and without trisomy 21, we noticed many heat shock genes are up-regulated in blood samples from individuals with trisomy 21. Yet no major heat shock response regulating transcription factor is encoded on chromosome 21, leaving it unclear why trisomy 21 itself would cause a heat shock response, or how it would impact the ability of blood cells to subsequently respond when faced with heat shock stress. To explore these issues in a context independent of any trisomy 21 associated co-morbidities or developmental differences, we characterized the response to heat shock of two lymphoblastoid cell lines derived from brothers with and without trisomy 21. To carefully compare the chromatin state and the transcription status of these cell lines, we measured nascent transcription, chromatin accessibility, and single cell transcript levels in the lymphoblastoid cell lines before and after acute heat shock treatment. The trisomy 21 cells displayed a more robust heat shock response after just one hour at 42{degrees}C than the matched disomic cells. We suggest multiple potential mechanisms for this increased heat shock response in lymphoblastoid cells with trisomy 21 including the possibility that cells with trisomy 21 may exist in a hyper-reactive state due to chronic stresses. Whatever the mechanism, abnormal heat shock response in individuals with Down syndrome may hobble immune responses during fever and contribute to health problems in these individuals.

bioinformatics↗

bioRxiv2021

Myonuclear maturation dynamics in aged and adult regenerating mouse skeletal muscle

Skeletal muscle cells are multinucleated syncytial cells arising from cell fusion, yet despite sharing a common cytoplasm individual myonuclei express distinct transcriptional programs. Whether individual myonuclei acquire heterogenous transcriptional states via differences in their progenitors, during differentiation, or once their anatomical position is acquired, is not known. We performed transcriptome and pseudotime analysis of single myogenic nuclei from uninjured and post-injury murine skeletal muscle to assess when myonuclear heterogeneity is acquired. Two distinct progenitors contribute to myonuclei, one a non-myogenic fibroblast subtype, and skeletal muscle stem cells the other. Both progenitors enter a single pseudotime trajectory that bifurcates as myonuclei mature into two branches segregated by myosin isoform expression and metabolic profiles, suggesting transcriptional heterogeneity is acquired as myonuclei mature. In aged skeletal muscle myogenic progenitor expansion is perturbed and nuclei from aged muscle display distinct pseudotemporal kinetics compared to nuclei from young mice. In aged mice, the inferred myogenic differentiation trajectory is delayed, altering the distribution of myogenic nuclei in pseudotime, suggesting that altered transcriptional dynamics in nuclei in aged mice may drive age-associated muscle deficits and bias myonuclei towards acquiring oxidative metabolic profiles.

molecular biology↗

bioRxiv2021

Infection of immune competent macrophages expressing functional Slc11a1 alters global gene expression, regulation of metal ions, and infection outcomes

Nutritional immunity involves cellular and physiological responses to invading pathogens, such as limiting iron availability, increasing exposure to bactericidal copper, and manipulating zinc to restrict the growth of pathogens. Manipulation of zinc at the host-pathogen interface depends on both the pathogens identity and the nature of the host cell. Here we examine infection of bone marrow-derived macrophages from 129S6/SvEvTac mice by Salmonella Typhimurium. Unlike Balb/c and C57BL/6 mice, 129S6/SvEvTac mice possess a functional Slc11a1 (Nramp-1), a phagosomal transporter of divalent cations. We carried out global RNA sequencing upon treatment with live or heat-killed Salmonella at 2 Hrs and 18 Hrs post-infection and observed widespread changes in metal transport, metal-dependent, and metal homeostasis genes, suggesting significant remodeling of iron, copper, and zinc availability by host cells. Changes in host cell gene expression suggest infection increases cytosolic zinc while simultaneously limiting zinc within the phagosome. Using a genetically encoded sensor, we demonstrate that cytosolic labile zinc increases 36-fold 12 hrs post-infection. Further, manipulation of zinc in the media alters bacterial clearance and replication, with zinc depletion inhibiting both processes. Comparing our results to published data on infection of C57BL/6 macrophages revealed notable differences in metal regulation and the global immune response, with 129S6 macrophages transitioning from M1 to M2 polarization over the course of infection and showing signs of recovery. Our results reveal that functional Slc11a1 profoundly affects the transcriptional landscape upon infection. Further, our results indicate that manipulation of zinc at the host-pathogen interface is more nuanced than that of iron or copper. 129S6 macrophage leverage intricate means of manipulating zinc availability and distribution to limit the pathogens access to zinc while simultaneously ensuring sufficient zinc to support the immune response. Author summaryMetal ions play an important role in influencing how immune cells such as macrophages respond to infection by pathogens. Because metal ions are both essential to survival, as well toxic when present is excessive amounts, the host and the pathogen have evolved diverse strategies to regulate metal acquisition and availability. Here, we show that the metal transporter slc11a1 plays a critical role in defining the host response to Salmonella infection. Infection causes widespread changes in expression of metal regulatory genes to limit the pathogens access to iron, increase its exposure to copper, and remodel zinc to ensure increased zinc in the cytosol and limited zinc for the pathogen. Macrophages expressing functional slc11a1 have a different profile of metal regulation and vastly different outcomes compared to immune compromised macrophage, demonstrating significantly different nutritional immune responses in immune competent versus immune compromised macrophages.

cell biology↗

bioRxiv2020

Ecology and molecular targets of hypermutation in the global microbiome.

Changes in the sequence of an organisms genome, i.e. mutations, are the raw material of evolution1. The frequency and location of mutations can be constrained by specific molecular mechanisms, such as Diversity-generating retroelements (DGRs)2-4. DGRs introduce mutations in specific target genes, and were characterized from several cultivated bacteria and bacteriophages2. Whilst a larger diversity of DGR loci has been identified in genomic data from environmental samples, i.e. metagenomes, the ecological role of these DGRs and their associated evolutionary drivers remain poorly understood5-7. Here we built and analyzed an extensive dataset of >30,000 metagenome-derived DGRs, and determine that DGRs have a single evolutionary origin and a universal bias towards adenine mutations. We further identified six major lineages of DGRs, each associated with a specific ecological niche defined as a genome type, i.e. whether the DGR is encoded on a viral or cellular genome, a limited set of taxa and environments, and a distinct type of target. Finally, we leverage read mapping and metagenomic time series to demonstrate that DGRs are consistently and broadly active, and responsible for >10% of all amino acid changes in some organisms at a conservative estimate. Overall, these results highlight the strong constraints under which DGRs diversify and expand, and elucidate several distinct roles these elements play in natural communities and in shaping microbial community structure and function in our environment.

microbiology↗

bioRxiv2019

Nascent transcript analysis of glucocorticoid crosstalk with TNF defines primary and cooperative inflammatory repression

The glucocorticoid receptor (GR) binds to specific DNA sequences and directly induces transcription of anti-inflammatory genes that contribute to cytokine repression, frequently in cooperation with NF-kB. Whether inflammatory repression also occurs through local interactions between GR and inflammatory gene regulatory elements remains controversial. Here, using Global Run-on Sequencing (GRO-seq) in human airway epithelial cells, we show that glucocorticoid signaling represses transcription within 10 minutes. Many repressed regulatory regions reside within hyper-ChIPable genomic regions that are subject to non-specific interactions with some antibodies. When this was accounted for, we determined that transcriptional repression occurs without local GR occupancy. Instead, widespread transcriptional induction through canonical GR binding sites is associated with reciprocal repression of distal TNF-regulated enhancers through a chromatin-dependent process, as evidenced by chromatin accessibility and enhancer-reporter assays. Simultaneously, transcriptional induction of key anti-inflammatory effectors is decoupled from primary repression through cooperation between GR and NF-kB at a subset of regulatory regions. Thus, glucocorticoids exert bimodal restraints on inflammation characterized by rapid primary transcriptional repression without local GR occupancy and secondary anti-inflammatory effects resulting from transcriptional cooperation between GR and NF-kB.

molecular biology↗