Daniel Richard

All Publications

• Targeting immune cells in the aged brain reveals that engineered cytokine IL-10 enhances neurogenesis and improves cognition. Immunity Navarro Negredo, P., You, J., Hauptschein, M., Schroer, A. B., Richard, D. J., Abhiraman, G. C., Tsai, A. P., Sun, E. D., Notarangelo, G., Ramirez-Matias, J., Zhou, O. Y., Buckley, M. T., Malacon, K. E., Xu, L., Sucharov, J., Ramirez Lopez, E., Picton, L., Wyss-Coray, T., Saxton, R. A., Fernandes, R. A., Villeda, S. A., Garcia, K. C., Brunet, A. 2026

  Abstract

  The immune system could play an important role in the age-related decline in brain function, yet specific immune-based strategies to enhance brain resilience in older individuals are lacking. Here, we combined engineered proteins and direct brain delivery to target immune cell populations within the old brain. We detected T cells with an exhaustion signature in the old brain and targeted them with a potent engineered checkpoint inhibitor (RIPR-PD1). This led to T cell expansion and strong pro-inflammatory responses in many brain cell types, notably microglia. To rescue age-related inflammatory imbalances in microglia, we used the anti-inflammatory cytokine interleukin (IL)-10. IL-10 boosted anti-inflammatory responses in old microglia, but it also triggered pro-inflammatory signaling. An engineered IL-10 variant that uncouples pro- and anti-inflammatory responses positively impacted the transcriptome of multiple cell types, enhanced neurogenesis, and improved cognition in aged mice. Our findings pave the way for immunotherapies for the aged brain.

  View details for DOI 10.1016/j.immuni.2026.01.016

  View details for PubMedID 41619730

• Age-related microbiome metabolites alter RNA splicing and chromatin accessibility in the brain. bioRxiv : the preprint server for biology Chakraborty, M., Shi, S. M., Porter, I. E., Richard, D. J., Marinov, G. K., Moore, A. A., Blum, J. L., Natarajan, A., Jahng, J. W., Wu, J. C., Lu, S. X., Davidson, S. M., Greenleaf, W. J., Saw, N. L., Shamloo, M., Brunet, A., Wyss-Coray, T., Bhatt, A. S. 2025

  Abstract

  The gut microbiome generates diverse metabolites that can enter the bloodstream and alter host biology, including brain function. Hundreds of physiologically relevant, gut-brain signaling molecules likely exist; however, there has been no systematic, high-throughput effort to identify and validate them. Here, we integrate computational, in vitro, and in vivo approaches to pinpoint microbiome-derived metabolites whose blood levels change during aging, and that induce corresponding changes in the mouse brain. First, we mine large-scale metabolomics datasets from human cohorts (each n ≥ 1200) to identify 30 microbiome-associated metabolites whose blood levels change with age. We then screen this panel in an in vitro transcriptomic assay to identify metabolites that perturb genes linked to age-related neurodegeneration. We then test four metabolites in an acute-exposure mouse model, and use multi-omic approaches to evaluate their impact on cellular functions in the brain. We confirm the known neurodegeneration-promoting effects of trimethylamine N-oxide (TMAO), including mitochondrial dysfunction, and further discover its disruptive impact on the pathways of glycolysis, GABAergic signaling, and RNA splicing. Additionally, we identify glycodeoxycholic acid (GDCA), a microbiome-derived secondary bile acid, as a potent regulator of chromatin accessibility and suppressor of genes that protect the brain from age-related, neurodegeneration-promoting insults. GDCA also acutely reduces mobility. In summary, we present a scalable framework for linking microbiome metabolites to host pathologies, and apply it to identify microbial metabolites that induce molecular changes related to neurodegeneration.

  View details for DOI 10.1101/2025.10.03.680371

  View details for PubMedID 41256397

  View details for PubMedCentralID PMC12621679

• The evolution of hominin bipedalism in two steps. Nature Senevirathne, G., Fernandopulle, S. C., Richard, D., Baumgart, S. L., Christensen, A. L., Fabbri, M., Höppner, J., Jüppner, H., Li, P., Bothe, V., Fröbisch, N., Simcock, I., Arthurs, O. J., Calder, A., Freilich, N., Nowlan, N. C., Glass, I. A., Craft, A., Capellini, T. D. 2025

  Abstract

  Bipedalism is a human-defining trait1-3. It is made possible by the familiar, bowl-shaped pelvis, whose short, wide iliac blades curve along the sides of the body to stabilize walking and support internal organs and a large-brained, broad-shouldered baby4-6. The ilium changes compared with living primates are an evolutionary novelty7. However, how this evolution came about remains unknown. Here, using a multifaceted histological, comparative genomic and functional genomic approach, we identified the developmental bases of the morphogenetic shifts in the human pelvis that made bipedalism possible. First, we observe that the human ilium cartilage growth plate underwent a heterotopic shift, residing perpendicular to the orientation present in other primate (and mouse) ilia. Second, we observe heterochronic and heterotopic shifts in ossification that are unlike those in non-human primate ilia or human long bones. Ossification initiates posteriorly, resides externally with fibroblast (and perichondral) cells contributing to osteoblasts, and is delayed compared with other bones in humans and with primate ilia. Underlying these two shifts are regulatory changes in an integrated chondrocyte-perichondral-osteoblast pathway, involving complex hierarchical interactions between SOX9-ZNF521-PTH1R and RUNX2-FOXP1/2. These innovations facilitated further growth of the human pelvis and the unique formation of the ilium among primates.

  View details for DOI 10.1038/s41586-025-09399-9

  View details for PubMedID 40866708

  View details for PubMedCentralID 6628783

• Complex regulatory interactions at GDF5 shape joint morphology and osteoarthritis disease risk. Arthritis & rheumatology (Hoboken, N.J.) Coveney, C. R., Maridas, D., Chen, H., Muthuirulan, P., Liu, Z., Jagoda, E., Yarlagadda, S., Movahhedi, M., Proffen, B., Dashtdar, B., Aghaalikhani, M., Richard, D., Rosen, V., Kiapour, A. M., Capellini, T. D. 2025

  Abstract

  To reveal causal level osteoarthritis (OA) disease biology by targeting regulatory interactions at GDF5.By investigating different GDF5 regulatory regions (R2, R3-5, R7-R9, R18-20, GROW1) we explored their functional impacts on gene expression and joint morphology in vivo and in vitro. We additionally modeled OA variants in said enhancers in vitro and in vivo mouse models for expression and disease effects.For all regulatory regions we found evidence of activation/repression between or within said regions that impacted patterns of joint-specific expression. Examples are: (1) the R4 enhancer, whilst considered to be activating, has dual roles repressing expression in adjacent tissues and sites; and (2) Growth plate-specific expression patterns by the GROW1 regulatory region are confined by adjacent sequences to restrict its expression to the perichondrium. We next targeted different regions/variants in vivo. Testing the R2de region resulted in ~40% reduction in Gdf5 expression, joint morphology changes, but no increase in OA risk; likewise, modeling the most cited OA risk (rs143384) variant in mice had no impact on expression, joint morphology, or disease. However, we identified epistatic interactions between this rs143384 risk variant and downstream disease risk variants lying within regulatory regions subject to repression, that compound to impact expression.These findings, at the best studied OA locus to date, serve as lessons on the nature of how gene regulatory interactions and local epistasis work in the etiology of OA disease risk, and that assessment of individual variants of high GWAS significance need not alone be considered causal.

  View details for DOI 10.1002/art.43231

  View details for PubMedID 40356240

• Functional genomics of human skeletal development and the patterning of height heritability CELL Richard, D., Muthuirulan, P., Young, M., Yengo, L., Vedantam, S., Marouli, E., Bartell, E., GIANT Consortium, G. T., Hirschhorn, J., Capellini, T. D. 2025; 188 (1): 15-32.e24

  Abstract

  Underlying variation in height are regulatory changes to chondrocytes, cartilage cells comprising long-bone growth plates. Currently, we lack knowledge on epigenetic regulation and gene expression of chondrocytes sampled across the human skeleton, and therefore we cannot understand basic regulatory mechanisms controlling height biology. We first rectify this issue by generating extensive epigenetic and transcriptomic maps from chondrocytes sampled from different growth plates across developing human skeletons, discovering novel regulatory networks shaping human bone/joint development. Next, using these maps in tandem with height genome-wide association study (GWAS) signals, we disentangle the regulatory impacts that skeletal element-specific versus global-acting variants have on skeletal growth, revealing the prime importance of regulatory pleiotropy in controlling height variation. Finally, as height is highly heritable, and thus often the test case for complex-trait genetics, we leverage these datasets within a testable omnigenic model framework to discover novel chondrocyte developmental modules and peripheral-acting factors shaping height biology and skeletal growth.

  View details for DOI 10.1016/j.cell.2024.10.040

  View details for Web of Science ID 001412813600001

  View details for PubMedID 39549696

  View details for PubMedCentralID PMC11724752

• Disagreement on foundational principles of biological aging. PNAS nexus Gladyshev, V. N., Anderson, B., Barlit, H., Barré, B., Beck, S., Behrouz, B., Belsky, D. W., Chaix, A., Chamoli, M., Chen, B. H., Cheng, K., Chuprin, J., Churchill, G. A., Cipriano, A., Colville, A., Deelen, J., Deigin, Y., Edmonds, K. K., English, B. W., Fang, R., Florea, M., Gershteyn, I. M., Gill, D., Goetz, L. H., Gorbunova, V., Griffin, P. T., Horvath, S., Borch Jensen, M., Jin, X., Jovanovska, S., Kajderowicz, K. M., Kasahara, T., Kerepesi, C., Kulkarni, S., Labunskyy, V. M., Levine, M. E., Libert, S., Lu, J. Y., Lu, Y. R., Marioni, R. E., McCoy, B. M., Mitchell, W., Moqri, M., Nasirian, F., Niimi, P., Oh, H. S., Okundaye, B., Parkhitko, A. A., Peshkin, L., Petljak, M., Poganik, J. R., Pridham, G., Promislow, D. E., Prusisz, W., Quiniou, M., Raj, K., Richard, D., Ricon, J. L., Rutledge, J., Scheibye-Knudsen, M., Schork, N. J., Seluanov, A., Shadpour, M., Shindyapina, A. V., Shuken, S. R., Sivakumar, S., Stoeger, T., Sugiura, A., Sutton, N. R., Suvorov, A., Tarkhov, A. E., Teeling, E. C., Trapp, A., Tyshkovskiy, A., Unfried, M., Ward-Caviness, C. K., Yim, S. H., Ying, K., Yunes, J., Zhang, B., Zhavoronkov, A. 2024; 3 (12): pgae499

  Abstract

  To gain insight into how researchers of aging perceive the process they study, we conducted a survey among experts in the field. While highlighting some common features of aging, the survey exposed broad disagreement on the foundational issues. What is aging? What causes it? When does it begin? What constitutes rejuvenation? Not only was there no consensus on these and other core questions, but none of the questions received a majority opinion-even regarding the need for consensus itself. Despite many researchers believing they understand aging, their understanding diverges considerably. Importantly, as different processes are labeled as "aging" by researchers, different experimental approaches are prioritized. The survey shed light on the need to better define which aging processes this field should target and what its goals are. It also allowed us to categorize contemporary views on aging and rejuvenation, revealing critical, yet largely unanswered, questions that appear disconnected from the current research focus. Finally, we discuss ways to address the disagreement, which we hope will ultimately aid progress in the field.

  View details for DOI 10.1093/pnasnexus/pgae499

  View details for PubMedID 39660064

  View details for PubMedCentralID PMC11630784

• Lineage-specific differences and regulatory networks governing human chondrocyte development ELIFE Richard, D., Pregizer, S., Venkatasubramanian, D., Raftery, R. M., Muthuirulan, P., Liu, Z., Capellini, T. D., Craft, A. M. 2023; 12

  Abstract

  To address large gaps in our understanding of the molecular regulation of articular and growth plate cartilage development in humans, we used our directed differentiation approach to generate these distinct cartilage tissues from human embryonic stem cells. The resulting transcriptomic profiles of hESC-derived articular and growth plate chondrocytes were similar to fetal epiphyseal and growth plate chondrocytes, with respect to genes both known and previously unknown to cartilage biology. With the goal to characterize the regulatory landscapes accompanying these respective transcriptomes, we mapped chromatin accessibility in hESC-derived chondrocyte lineages, and mouse embryonic chondrocytes, using ATAC-sequencing. Integration of the expression dataset with the differentially accessible genomic regions revealed lineage-specific gene regulatory networks. We validated functional interactions of two transcription factors (TFs) (RUNX2 in growth plate chondrocytes and RELA in articular chondrocytes) with their predicted genomic targets. The maps we provide thus represent a framework for probing regulatory interactions governing chondrocyte differentiation. This work constitutes a substantial step towards comprehensive and comparative molecular characterizations of distinct chondrogenic lineages and sheds new light on human cartilage development and biology.

  View details for DOI 10.7554/eLife.79925

  View details for Web of Science ID 000964336400001

  View details for PubMedID 36920035

  View details for PubMedCentralID PMC10069868

• Regulatory dissection of the severe COVID-19 risk locus introgressed by Neanderthals ELIFE Jagoda, E., Marnetto, D., Senevirathne, G., Gonzalez, V., Baid, K., Montinaro, F., Richard, D., Falzarano, D., LeBlanc, E., Colpitts, C. C., Banerjee, A., Pagani, L., Capellini, T. D. 2023; 12

  Abstract

  Individuals infected with the SARS-CoV-2 virus present with a wide variety of symptoms ranging from asymptomatic to severe and even lethal outcomes. Past research has revealed a genetic haplotype on chromosome 3 that entered the human population via introgression from Neanderthals as the strongest genetic risk factor for the severe response to COVID-19. However, the specific variants along this introgressed haplotype that contribute to this risk and the biological mechanisms that are involved remain unclear. Here, we assess the variants present on the risk haplotype for their likelihood of driving the genetic predisposition to severe COVID-19 outcomes. We do this by first exploring their impact on the regulation of genes involved in COVID-19 infection using a variety of population genetics and functional genomics tools. We then perform a locus-specific massively parallel reporter assay to individually assess the regulatory potential of each allele on the haplotype in a multipotent immune-related cell line. We ultimately reduce the set of over 600 linked genetic variants to identify four introgressed alleles that are strong functional candidates for driving the association between this locus and severe COVID-19. Using reporter assays in the presence/absence of SARS-CoV-2, we find evidence that these variants respond to viral infection. These variants likely drive the locus' impact on severity by modulating the regulation of two critical chemokine receptor genes: CCR1 and CCR5. These alleles are ideal targets for future functional investigations into the interaction between host genomics and COVID-19 outcomes.

  View details for DOI 10.7554/eLife.71235

  View details for Web of Science ID 000932804200001

  View details for PubMedID 36763080

  View details for PubMedCentralID PMC9917435

• The developmental impacts of natural selection on human pelvic morphology SCIENCE ADVANCES Young, M., Richard, D., Grabowski, M., Auerbach, B. M., de Bakker, B. S., Hagoort, J., Muthuirulan, P., Kharkar, V., Kurki, H. K., Betti, L., Birkenstock, L., Lewton, K. L., Capellini, T. D. 2022; 8 (33): eabq4884

  Abstract

  Evolutionary responses to selection for bipedalism and childbirth have shaped the human pelvis, a structure that differs substantially from that in apes. Morphology related to these factors is present by birth, yet the developmental-genetic mechanisms governing pelvic shape remain largely unknown. Here, we pinpoint and characterize a key gestational window when human-specific pelvic morphology becomes recognizable, as the ilium and the entire pelvis acquire traits essential for human walking and birth. We next use functional genomics to molecularly characterize chondrocytes from different pelvic subelements during this window to reveal their developmental-genetic architectures. We then find notable evidence of ancient selection and genetic constraint on regulatory sequences involved in ilium expansion and growth, findings complemented by our phenotypic analyses showing that variation in iliac traits is reduced in humans compared to African apes. Our datasets provide important resources for musculoskeletal biology and begin to elucidate developmental mechanisms that shape human-specific morphology.

  View details for DOI 10.1126/sciadv.abq4884

  View details for Web of Science ID 000842064500038

  View details for PubMedID 35977020

  View details for PubMedCentralID PMC9385149

• Intronic regulation of SARS-CoV-2 receptor (ACE2) expression mediated by immune signaling and oxidative stress pathways ISCIENCE Richard, D., Muthuirulan, P., Aguiar, J., Doxey, A. C., Banerjee, A., Mossman, K., Hirota, J., Capellini, T. D. 2022; 25 (7): 104614

  Abstract

  The angiotensin-converting enzyme 2 (ACE2) protein is a key catalytic regulator of the renin-angiotensin system (RAS), involved in fluid homeostasis and blood pressure modulation. ACE2 also serves as a cell-surface receptor for some coronaviruses such as SARS-CoV and SARS-CoV-2. Improved characterization of ACE2 regulation may help us understand the effects of pre-existing conditions on COVID-19 incidence, as well as pathogenic dysregulation following viral infection. Here, we perform bioinformatic analyses to hypothesize on ACE2 gene regulation in two different physiological contexts, identifying putative regulatory elements of ACE2 expression. We perform functional validation of our computational predictions via targeted CRISPR-Cas9 deletions of these elements in vitro, finding them responsive to immune signaling and oxidative-stress pathways. This contributes to our understanding of ACE2 gene regulation at baseline and immune challenge. Our work supports pursuit of these putative mechanisms in our understanding of infection/disease caused by current, and future, SARS-related viruses such as SARS-CoV-2.

  View details for DOI 10.1016/j.isci.2022.104614

  View details for Web of Science ID 000825198600008

  View details for PubMedID 35756893

  View details for PubMedCentralID PMC9213013

• The active grandparent hypothesis: Physical activity and the evolution of extended human healthspans and lifespans PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Lieberman, D. E., Kistner, T. M., Richard, D., Lee, I., Baggish, A. L. 2021; 118 (50)

  Abstract

  The proximate mechanisms by which physical activity (PA) slows senescence and decreases morbidity and mortality have been extensively documented. However, we lack an ultimate, evolutionary explanation for why lifelong PA, particularly during middle and older age, promotes health. As the growing worldwide epidemic of physical inactivity accelerates the prevalence of noncommunicable diseases among aging populations, integrating evolutionary and biomedical perspectives can foster new insights into how and why lifelong PA helps preserve health and extend lifespans. Building on previous life-history research, we assess the evidence that humans were selected not just to live several decades after they cease reproducing but also to be moderately physically active during those postreproductive years. We next review the longstanding hypothesis that PA promotes health by allocating energy away from potentially harmful overinvestments in fat storage and reproductive tissues and propose the novel hypothesis that PA also stimulates energy allocation toward repair and maintenance processes. We hypothesize that selection in humans for lifelong PA, including during postreproductive years to provision offspring, promoted selection for both energy allocation pathways which synergistically slow senescence and reduce vulnerability to many forms of chronic diseases. As a result, extended human healthspans and lifespans are both a cause and an effect of habitual PA, helping explain why lack of lifelong PA in humans can increase disease risk and reduce longevity.

  View details for DOI 10.1073/pnas.2107621118

  View details for Web of Science ID 000732715700022

  View details for PubMedID 34810239

  View details for PubMedCentralID PMC8685690

• GWAS meta-analysis followed by Mendelian randomization revealed potential control mechanisms for circulating α-Klotho levels HUMAN MOLECULAR GENETICS Gergei, I., Zheng, J., Andlauer, T. F. M., Brandenburg, V., Mirza-Schreiber, N., Mueller-Myhsok, B., Kraemer, B. K., Richard, D., Falk, L., Moverare-Skrtic, S., Ohlsson, C., Smith, G., Maerz, W., Voelkl, J., Tobias, J. H. 2022; 31 (5): 792-802

  Abstract

  The protein α-Klotho acts as transmembrane co-receptor for fibroblast growth factor 23 (FGF23) and is a key regulator of phosphate homeostasis. However, α-Klotho also exists in a circulating form, with pleiotropic, but incompletely understood functions and regulation. Therefore, we undertook a genome-wide association study (GWAS) meta-analysis followed by Mendelian randomization (MR) of circulating α-Klotho levels. Plasma α-Klotho levels were measured by enzyme-linked immunosorbent assay (ELISA) in the Ludwigshafen Risk and Cardiovascular Health and Avon Longitudinal Study of Parents and Children (mothers) cohorts, followed by a GWAS meta-analysis in 4376 individuals across the two cohorts. Six signals at five loci were associated with circulating α-Klotho levels at genome-wide significance (P < 5 × 10-8), namely ABO, KL, FGFR1, and two post-translational modification genes, B4GALNT3 and CHST9. Together, these loci explained >9% of the variation in circulating α-Klotho levels. MR analyses revealed no causal relationships between α-Klotho and renal function, FGF23-dependent factors such as vitamin D and phosphate levels, or bone mineral density. The screening for genetic correlations with other phenotypes followed by targeted MR suggested causal effects of liability of Crohn's disease risk [Inverse variance weighted (IVW) beta = 0.059 (95% confidence interval 0.026, 0.093)] and low-density lipoprotein cholesterol levels [-0.198 (-0.332, -0.063)] on α-Klotho. Our GWAS findings suggest that two enzymes involved in post-translational modification, B4GALNT3 and CHST9, contribute to genetic influences on α-Klotho levels, presumably by affecting protein turnover and stability. Subsequent evidence from MR analyses on α-Klotho levels suggest regulation by mechanisms besides phosphate-homeostasis and raise the possibility of cross-talk with FGF19- and FGF21-dependent pathways, respectively. Significance statement: α-Klotho as a transmembrane protein is well investigated along the endocrine FGF23-α-Klotho pathway. However, the role of the circulating form of α-Klotho, which is generated by cleavage of transmembrane α-Klotho, remains incompletely understood. Genetic analyses might help to elucidate novel regulatory and functional mechanisms. The identification of genetic factors related to circulating α-Klotho further enables MR to examine causal relationships with other factors. The findings from the first GWAS meta-analysis of circulating α-Klotho levels identified six genome-wide significant signals across five genes. Given the function of two of the genes identified, B4GALNT3 and CHST9, it is tempting to speculate that post-translational modification significantly contributes to genetic influences on α-Klotho levels, presumably by affecting protein turnover and stability.

  View details for DOI 10.1093/hmg/ddab263

  View details for Web of Science ID 000756570900001

  View details for PubMedID 34542150

  View details for PubMedCentralID PMC8895756

• Joint disease-specificity at the regulatory base-pair level NATURE COMMUNICATIONS Muthuirulan, P., Zhao, D., Young, M., Richard, D., Liu, Z., Emami, A., Portilla, G., Hosseinzadeh, S., Cao, J., Maridas, D., Sedlak, M., Menghini, D., Cheng, L., Li, L., Ding, X., Ding, Y., Rosen, V., Kiapour, A. M., Capellini, T. D. 2021; 12 (1): 4161

  Abstract

  Given the pleiotropic nature of coding sequences and that many loci exhibit multiple disease associations, it is within non-coding sequence that disease-specificity likely exists. Here, we focus on joint disorders, finding among replicated loci, that GDF5 exhibits over twenty distinct associations, and we identify causal variants for two of its strongest associations, hip dysplasia and knee osteoarthritis. By mapping regulatory regions in joint chondrocytes, we pinpoint two variants (rs4911178; rs6060369), on the same risk haplotype, which reside in anatomical site-specific enhancers. We show that both variants have clinical relevance, impacting disease by altering morphology. By modeling each variant in humanized mice, we observe joint-specific response, correlating with GDF5 expression. Thus, we uncouple separate regulatory variants on a common risk haplotype that cause joint-specific disease. By broadening our perspective, we finally find that patterns of modularity at GDF5 are also found at over three-quarters of loci with multiple GWAS disease associations.

  View details for DOI 10.1038/s41467-021-24345-9

  View details for Web of Science ID 000672715200007

  View details for PubMedID 34230488

  View details for PubMedCentralID PMC8260791

• Shifting epigenetic contexts influence regulatory variation and disease risk AGING-US Richard, D., Capellini, T. D. 2021; 13 (12): 15699-15749

  Abstract

  Epigenetic shifts are a hallmark of aging that impact transcriptional networks at regulatory level. These shifts may modify the effects of genetic regulatory variants during aging and contribute to disease pathomechanism. However, these shifts occur on the backdrop of epigenetic changes experienced throughout an individual's development into adulthood; thus, the phenotypic, and ultimately fitness, effects of regulatory variants subject to developmental- versus aging-related epigenetic shifts may differ considerably. Natural selection therefore may act differently on variants depending on their changing epigenetic context, which we propose as a novel lens through which to consider regulatory sequence evolution and phenotypic effects. Here, we define genomic regions subjected to altered chromatin accessibility as tissues transition from their fetal to adult forms, and subsequently from early to late adulthood. Based on these epigenomic datasets, we examine patterns of evolutionary constraint and potential functional impacts of sequence variation (e.g., genetic disease risk associations). We find that while the signals observed with developmental epigenetic changes are consistent with stronger fitness consequences (i.e., negative selection pressures), they tend to have weaker effects on genetic risk associations for aging-related diseases. Conversely, we see stronger effects of variants with increased local accessibility in adult tissues, strongest in young adult when compared to old. We propose a model for how epigenetic status of a region may influence the effects of evolutionary relevant sequence variation, and suggest that such a perspective on gene regulatory networks may elucidate our understanding of aging biology.

  View details for Web of Science ID 000669027700003

  View details for PubMedID 34138751

  View details for PubMedCentralID PMC8266365

• Experimental and natural evidence of SARS-CoV-2-infection-induced activation of type I interferon responses ISCIENCE Banerjee, A., El-Sayes, N., Budylowski, P., Jacob, R., Richard, D., Maan, H., Aguiar, J. A., Demian, W. L., Baid, K., D'Agostino, M. R., Ang, J., Murdza, T., Tremblay, B., Afkhami, S., Karimzadeh, M., Irving, A. T., Yip, L., Ostrowski, M., Hirota, J. A., Kozak, R., Capellini, T. D., Miller, M. S., Wang, B., Mubareka, S., McGeer, A. J., McArthur, A. G., Doxey, A. C., Mossman, K. 2021; 24 (5): 102477

  Abstract

  Type I interferons (IFNs) are our first line of defense against virus infection. Recent studies have suggested the ability of SARS-CoV-2 proteins to inhibit IFN responses. Emerging data also suggest that timing and extent of IFN production is associated with manifestation of COVID-19 severity. In spite of progress in understanding how SARS-CoV-2 activates antiviral responses, mechanistic studies into wild-type SARS-CoV-2-mediated induction and inhibition of human type I IFN responses are scarce. Here we demonstrate that SARS-CoV-2 infection induces a type I IFN response in vitro and in moderate cases of COVID-19. In vitro stimulation of type I IFN expression and signaling in human airway epithelial cells is associated with activation of canonical transcriptions factors, and SARS-CoV-2 is unable to inhibit exogenous induction of these responses. Furthermore, we show that physiological levels of IFNα detected in patients with moderate COVID-19 is sufficient to suppress SARS-CoV-2 replication in human airway cells.

  View details for DOI 10.1016/j.isci.2021.102477

  View details for Web of Science ID 000653990500086

  View details for PubMedID 33937724

  View details for PubMedCentralID PMC8074517

• Evolutionary Selection and Constraint on Human Knee Chondrocyte Regulation Impacts Osteoarthritis Risk CELL Richard, D., Liu, Z., Cao, J., Kiapour, A. M., Willen, J., Yarlagadda, S., Jagoda, E., Kolachalama, V. B., Sieker, J. T., Chang, G. H., Muthuirulan, P., Young, M., Masson, A., Konrad, J., Hosseinzadeh, S., Maridas, D. E., Rosen, V., Krawetz, R., Roach, N., Capellini, T. D. 2020; 181 (2): 362-+

  Abstract

  During human evolution, the knee adapted to the biomechanical demands of bipedalism by altering chondrocyte developmental programs. This adaptive process was likely not without deleterious consequences to health. Today, osteoarthritis occurs in 250 million people, with risk variants enriched in non-coding sequences near chondrocyte genes, loci that likely became optimized during knee evolution. We explore this relationship by epigenetically profiling joint chondrocytes, revealing ancient selection and recent constraint and drift on knee regulatory elements, which also overlap osteoarthritis variants that contribute to disease heritability by tending to modify constrained functional sequence. We propose a model whereby genetic violations to regulatory constraint, tolerated during knee development, lead to adult pathology. In support, we discover a causal enhancer variant (rs6060369) present in billions of people at a risk locus (GDF5-UQCC1), showing how it impacts mouse knee-shape and osteoarthritis. Overall, our methods link an evolutionarily novel aspect of human anatomy to its pathogenesis.

  View details for DOI 10.1016/j.cell.2020.02.057

  View details for Web of Science ID 000526184900017

  View details for PubMedID 32220312

  View details for PubMedCentralID PMC7179902

• Cataloging the Regulatory Landscape of Human Skeletal Development Via Functional Genomics: Insights into Human Pelvic Evolution Capellini, T. D., Young, M., Muthuirulan, P., Kharkar, V., Richard, D. WILEY. 2020: 43

  View details for Web of Science ID 000513288900164

• Using a Mouse Model to Study the Evolution of a Bipedal Trait: Characterizing the Regulatory Landscape of Muscles with Divergent MHC I Expression Queeno, S. R., Young, M., Richard, D., O'Neill, M. C., Capellini, T. D., Sterner, K. N. WILEY. 2019: 196

  View details for Web of Science ID 000458409602205

• Regulatory Constraint During Human Knee Evolution Underlies Osteoarthritis Risk Capellini, T. D., Richard, D., Jagoda, E. WILEY. 2019: 35

  View details for Web of Science ID 000458409600134

• Identification of Novel Loci Associated With Hip Shape: A Meta-Analysis of Genomewide Association Studies JOURNAL OF BONE AND MINERAL RESEARCH Baird, D. A., Evans, D. S., Kamanu, F. K., Gregory, J. S., Saunders, F. R., Giuraniuc, C. V., Barr, R. J., Aspden, R. M., Jenkins, D., Kiel, D. P., Orwoll, E. S., Cummings, S. R., Lane, N. E., Mullin, B. H., Williams, F. M. K., Richards, J., Wilson, S. G., Spector, T. D., Faber, B. G., Lawlor, D. A., Grundberg, E., Ohlsson, C., Pettersson-Kymmer, U., Capellini, T. D., Richard, D., Beck, T. J., Evans, D. M., Paternoster, L., Karasik, D., Tobias, J. H. 2019; 34 (2): 241-251

  Abstract

  We aimed to report the first genomewide association study (GWAS) meta-analysis of dual-energy X-ray absorptiometry (DXA)-derived hip shape, which is thought to be related to the risk of both hip osteoarthritis and hip fracture. Ten hip shape modes (HSMs) were derived by statistical shape modeling using SHAPE software, from hip DXA scans in the Avon Longitudinal Study of Parents and Children (ALSPAC; adult females), TwinsUK (mixed sex), Framingham Osteoporosis Study (FOS; mixed), Osteoporotic Fractures in Men study (MrOS), and Study of Osteoporotic Fractures (SOF; females) (total N = 15,934). Associations were adjusted for age, sex, and ancestry. Five genomewide significant (p < 5 × 10-9 , adjusted for 10 independent outcomes) single-nucleotide polymorphisms (SNPs) were associated with HSM1, and three SNPs with HSM2. One SNP, in high linkage disequilibrium with rs2158915 associated with HSM1, was associated with HSM5 at genomewide significance. In a look-up of previous GWASs, three of the identified SNPs were associated with hip osteoarthritis, one with hip fracture, and five with height. Seven SNPs were within 200 kb of genes involved in endochondral bone formation, namely SOX9, PTHrP, RUNX1, NKX3-2, FGFR4, DICER1, and HHIP. The SNP adjacent to DICER1 also showed osteoblast cis-regulatory activity of GSC, in which mutations have previously been reported to cause hip dysplasia. For three of the lead SNPs, SNPs in high LD (r2  > 0.5) were identified, which intersected with open chromatin sites as detected by ATAC-seq performed on embryonic mouse proximal femora. In conclusion, we identified eight SNPs independently associated with hip shape, most of which were associated with height and/or mapped close to endochondral bone formation genes, consistent with a contribution of processes involved in limb growth to hip shape and pathological sequelae. These findings raise the possibility that genetic studies of hip shape might help in understanding potential pathways involved in hip osteoarthritis and hip fracture. © 2018 The Authors. Journal of Bone and Mineral Research Published by Wiley Periodicals, Inc.

  View details for DOI 10.1002/jbmr.3605

  View details for Web of Science ID 000458653600005

  View details for PubMedID 30320955

  View details for PubMedCentralID PMC6375741

• Characterizing the regulatory landscape of human skeletal muscle tissue Queeno, S. R., O'Neill, M. C., Richard, D., Capellini, T. D., Sterner, K. N. WILEY. 2018: 216

  View details for Web of Science ID 000430656803121

• Epigenetic profiling of growth plate chondrocytes sheds insight into regulatory genetic variation influencing height ELIFE Guo, M., Liu, Z., Willen, J., Shaw, C. P., Richard, D., Jagoda, E., Doxey, A. C., Hirschhorn, J., Capellini, T. D. 2017; 6

  Abstract

  GWAS have identified hundreds of height-associated loci. However, determining causal mechanisms is challenging, especially since height-relevant tissues (e.g. growth plates) are difficult to study. To uncover mechanisms by which height GWAS variants function, we performed epigenetic profiling of murine femoral growth plates. The profiled open chromatin regions recapitulate known chondrocyte and skeletal biology, are enriched at height GWAS loci, particularly near differentially expressed growth plate genes, and enriched for binding motifs of transcription factors with roles in chondrocyte biology. At specific loci, our analyses identified compelling mechanisms for GWAS variants. For example, at CHSY1, we identified a candidate causal variant (rs9920291) overlapping an open chromatin region. Reporter assays demonstrated that rs9920291 shows allelic regulatory activity, and CRISPR/Cas9 targeting of human chondrocytes demonstrates that the region regulates CHSY1 expression. Thus, integrating biologically relevant epigenetic information (here, from growth plates) with genetic association results can identify biological mechanisms important for human growth.

  View details for DOI 10.7554/eLife.29329

  View details for Web of Science ID 000417121600001

  View details for PubMedID 29205154

  View details for PubMedCentralID PMC5716665

• Rainbow trout exposed to benzo[a]pyrene yields conserved microRNA binding sites in DNA methyltransferases across 500 million years of evolution SCIENTIFIC REPORTS Kuc, C., Richard, D. J., Johnson, S., Bragg, L., Servos, M. R., Doxey, A. C., Craig, P. M. 2017; 7: 16843

  Abstract

  The objective of this study was to examine the regulation of DNA methylation following acute (24 h) and prolonged (14 d) exposure to low (1 ng/L) and high (10 ng/L) benzo[a]pyrene. However, with the recent release of the rainbow trout genome, we were able to conduct a more detailed analysis regarding the regulation of the enzymes involved in DNA methylation; DNA methyltransferases (DNMTs). Bioinformatic approaches were used to identify candidate microRNA (miRNA) that potentially bind to the DNMT1 and DNMT3a 3'UTR. Results indicated a significant decrease in global methylation in both liver and muscle, with an associated decrease in DNA methyltransferase activity and DNMT3a transcript abundance. There was a significant increase in one specific candidate miRNA (miR29a) that was predicted to bind to DNMT3a. Taking a comparative genomics approach, the binding sites of miR29a to the DNMT3a 3'UTR was compared across species, spanning fish to mammals, and revealed a highly conserved binding motif that has been maintained since the vertebrate ancestor, approximately 500 million years ago. This research establishes that miRNA act as an essential mediator between the environment and DNA methylation patterns via DNMTs, which is further confirmed by a genomic regulatory mechanism that has been deeply conserved throughout evolution.

  View details for DOI 10.1038/s41598-017-17236-x

  View details for Web of Science ID 000417025400024

  View details for PubMedID 29203905

  View details for PubMedCentralID PMC5715007

• Lineage-specific mutational clustering in protein structures predicts evolutionary shifts in function BIOINFORMATICS Adams, J., Mansfield, M. J., Richard, D. J., Doxey, A. C. 2017; 33 (9): 1338-1345

  Abstract

  Spatially clustered mutations within specific regions of protein structure are thought to result from strong positive selection for altered protein functions and are a common feature of oncoproteins in cancer. Although previous studies have used spatial substitution clustering to identify positive selection between pairs of proteins, the ability of this approach to identify functional shifts in protein phylogenies has not been explored.We implemented a previous measure of spatial substitution clustering (the P3D statistic) and extended it to detect spatially clustered substitutions at specific branches of phylogenetic trees. We then applied the analysis to 423 690 phylogenetic branches from 9261 vertebrate protein families, and examined its ability to detect historical shifts in protein function. Our analysis identified 19 607 lineages from 5362 protein families in which substitutions were spatially clustered on protein structures at P3D  < 0.01. Spatially clustered substitutions were overrepresented among ligand-binding residues and were significantly enriched among particular protein families and functions including C2H2 transcription factors and protein kinases. A small but significant proportion of branches with spatially clustered substitution also were under positive selection according to the branch-site test. Lastly, exploration of the top-scoring candidates revealed historical substitution events in vertebrate protein families that have generated new functions and protein interactions, including ancient adaptations in SLC7A2, PTEN, and SNAP25 . Ultimately, our work shows that lineage-specific, spatially clustered substitutions are a useful feature for identifying functional shifts in protein families, and reveal new candidates for future experimental study.Source code and predictions for analyses performed in this study are available at: https://github.com/doxeylab/evoclust3d.acdoxey@uwaterloo.ca.Supplementary data are available at Bioinformatics online.

  View details for DOI 10.1093/bioinformatics/btw815

  View details for Web of Science ID 000402130100010

  View details for PubMedID 28052926