Eric Anthony Ortlund

Liver receptor homolog-1 (LRH-1; NR5A2) is a nuclear receptor that controls metabolic gene programs important for liver health. Here, we address two challenges with dissecting ligand-driven LRH-1 metabolism in vivo. First, we generate ME610, a potent LRH-1 agonist with favorable pharmacokinetic properties. Second, we engineer Nr5a2 ^humLBD mice, edited to recapitulate the ligand response of the human receptor. ME610 dose-dependently induces LRH-1 target genes and reduces triglycerides in livers of Nr5a2 ^humLBD mice. In a mouse model of diet-induced obesity (DIO), ME610 dramatically improves insulin sensitivity and reduces liver fat. Multi-omics integration of transcriptomics, lipidomics, and metabolomics data demonstrates a coordinated reversal of DIO-induced hepatic metabolic stress via changes in lipid, amino acid, and energy metabolism, with peroxisomal lipid metabolism emerging as a central process correlated to efficacy. These studies uncover new pathways through which LRH-1 regulates metabolism and demonstrate the therapeutic potential of synthetic LRH-1 agonists in obesity-associated metabolic diseases.

Rapid antigen tests (RATs) were a critical tool for diagnosing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection during the height of the pandemic, but their performance depends on antibodies that remain robust to viral evolution. Since most commercial RATs target the viral nucleocapsid protein, mutations in this domain can compromise binding and test performance. To systematically define this risk, we used deep mutational scanning (DMS) to map escape mutations for 82 antibodies across 43 commercially available tests, generating the most comprehensive escape database of SARS-CoV-2 viral mutations to date. Most antibodies target conformational epitopes, with 47 (57%) mapping to the RNA-binding domain and 23 (28%) to the dimerization domain. Principal component analysis and clustering revealed escape phenotypes, including shared features outside of predicted epitopes. As the pandemic proceeded, we noted a change in industry preference from IgG to IgM-type antibodies, which was associated with changes to epitope sub-class prevalence. Analysis of paired capture-detection antibodies revealed a preference for targeting the same structural domain (20/33 or 61%), indicating a slight preference for test antibodies to hit orthogonal yet proximal sites. Together, the results establish a comprehensive resource for understanding how SARS-CoV-2 nucleocapsid protein mutations affect antibody binding and the phenotypes that underlie RAT antibody selection. This resource and approach can help prioritize antibodies for follow-up performance testing and support the design of more mutation-resilient diagnostics. IMPORTANCE: It is critical during a pandemic for diagnostic tests to be accurate and for the false negative rate to be kept low. The rapidly changing nature of SARS-CoV-2 exposed a weakness with using rapid antigen tests to diagnose infection, as mutations to the viral nucleocapsid protein can cause the virus to evade detection. In this work, we systematically tested the robustness of 82 antibodies from 43 separate commercially available tests by deliberately mutating the nucleocapsid protein and investigating which changes caused weakened binding to the test antibodies. In addition to determining which specific tests were at risk from certain mutations, we collated our datasets into a meta-analysis, allowing us to identify some best practices for designing robust tests. We, therefore, expect that this resource will support the design of more mutation-resilient diagnostics, whether for future variants of SARS-CoV-2 or in applying these principles to other respiratory viruses, such as influenza virus.

The emergence of the SARS-CoV-2 Omicron BA.2.86 subvariant, a lineage derived from the BA.2 strain, led to the 2024–2025 COVID-19 vaccine update to include KP.2 or related JN.1-lineage spike antigens. We evaluated the magnitude, breadth, and durability of humoral immune responses following a single KP.2 vaccine dose in a longitudinal cohort of 21 individuals up to 6 months. KP.2 vaccination increased spike-specific binding and neutralizing antibodies against the ancestral WA.1 strain, as well as against the BA.5, XBB.1.5, and KP.2 variants. Power-law modeling estimated half-lives for WA.1- and KP.2-specific IgG responses at 770 and 248 days, respectively. Additionally, the KP.2 dose increased IgG1 and IgG4 subclasses more than IgG2 and IgG3 responses to both spike proteins. Serum-depletion experiments using WA.1 or KP.2 proteins demonstrated that most vaccine-elicited antibodies were cross-reactive. Consequently, KP.2 vaccine-induced antibodies retained broad neutralizing activity against recently circulating Omicron subvariants (BA.2.86, KP.3.1.1, XEC, LP.8.1, LF.7, XFG.3.12, PQ.1, BA.3.2.1, and RE.2). Using a live virus neutralization assay, XFG.3.12 showed the greatest reduction in neutralizing titers relative to KP.2 (4.2-fold). In a small subset, an LP.8.1 vaccine dose increased neutralizing activity against the matched variant while maintaining WA.1 and KP.2 cross-reactivity, but only modestly increased antibodies to divergent variants BA.3.2.1 and RE.2. Ultimately, these data indicate the KP.2 mRNA vaccine generates durable, cross-reactive responses against current Omicron subvariants. However, ongoing spike evolution impacts the neutralization of emerging lineages, highlighting the need for continued viral monitoring and timely vaccine updates. IMPORTANCE: SARS-CoV-2 continues to evolve, raising ongoing concerns about how well updated vaccines protect against emerging variants. This study evaluates antibody responses after a KP.2 spike mRNA vaccine dose. It shows that a single dose induces durable and broadly cross-reactive immunity against both earlier strains and recently circulating Omicron subvariants. Despite this breadth, reduced neutralizing activity against certain emerging variants indicates that ongoing antigenic changes can impact vaccine-induced antibody effectiveness. These findings provide insight into how current vaccines perform over time and highlight the need to track viral evolution and update vaccine antigens to maintain broad protection against severe disease, hospitalization, and death.

Acyl-CoA thioesterase 11 (ACOT11) regulates energy expenditure in brown adipose tissue by limiting availability of activated fatty acids to reach the mitochondria where they may be used as fuel. To balance energy demand with fuel conservation, ACOT11 is allosterically regulated by cellular lipid composition, sensed through its C-terminal steroidogenic acute regulatory transfer-related (START) domain, and by cellular energy status, mediated through the binding of ATP or ADP within the tandem thioesterase domains. Here, we present the first full-length cryo-EM structures of ACOT11 in CoA-, ADP-, and ATP-bound states. The CoA-bound structure defines the product-binding site and supports a catalytic mechanism involving Asp74, Thr91, and Asn232, with mutagenesis of Asp74 and Asn232 markedly reducing thioesterase activity. ADP binding orders a conserved regulatory loop and stabilizes the regulatory helix–START domain module, whereas ATP adopts protomer-specific conformations, including an “up and out” pose associated with reduced local order near the regulatory helix. Together, these data support a model in which ATP/ADP-dependent remodeling of a regulatory loop–helix module shifts ACOT11 between distinct regulatory states and provides a structural framework for how cellular energy status may be coupled to fatty acyl-CoA metabolism.

The emergence of the SARS-CoV-2 Omicron BA.2.86 subvariant, a lineage derived from the BA.2 strain, led to the 2024-2025 COVID-19 vaccine update to include KP.2 or related JN.1-lineage spike antigens. We evaluated the magnitude, breadth, and durability of humoral immune responses following a single KP.2 vaccine dose in a longitudinal cohort of 21 individuals up to six months. KP.2 vaccination increased spike-specific binding and neutralizing antibodies against the ancestral WA1 strain, alongside the BA.5, XBB.1.5, and KP.2 variants. Power law modeling estimated half-lives for WA1- and KP.2-specific IgG responses at 770 and 248 days, respectively. Additionally, the KP.2 dose increased IgG1 and IgG4 subclasses more than IgG2 and IgG3 responses to both spike proteins. Serum depletion experiments using WA1 or KP.2 proteins demonstrated most vaccine-elicited antibodies were cross-reactive. Consequently, KP.2 vaccine-induced antibodies retained broad neutralizing activity against recently circulating Omicron subvariants (BA.2.86, KP.3.1.1, XEC, LP.8.1, LF.7, XFG.3.12, PQ.1, BA.3.2.1, and RE.2). Using a live virus neutralization assay, XFG.3.12 showed the greatest reduction in neutralizing titers relative to KP.2 (4.2-fold). In a small subset, an LP.8.1 vaccine dose increased neutralizing activity against the matched variant while maintaining WA1 and KP.2 cross-reactivity, but only modestly increased antibodies to divergent variants BA.3.2.1 and RE.2. Ultimately, these data indicate the KP.2 mRNA vaccine generates durable, cross-reactive responses against current Omicron subvariants. However, ongoing spike evolution impacts neutralization of emerging lineages, highlighting the need for continued viral monitoring and timely vaccine updates. IMPORTANCE: SARS-CoV-2 continues to evolve, raising ongoing concerns about how well updated vaccines protect against emerging variants. This study evaluates antibody responses after KP.2 spike mRNA vaccine dose and shows that a single dose induces durable and broadly cross-reactive immunity against both earlier strains and recently circulating Omicron subvariants. Despite this breadth, reduced neutralizing activity against certain emerging variants indicates that ongoing antigenic changes can impact vaccine induced antibody effectiveness. These findings provide insight into how current vaccines perform over time and highlight the need to track viral evolution and update vaccine antigens to maintain broad protection against severe disease, hospitalization, and death.

Background: Exercise training is known to benefit health and reduce disease risk. While skeletal muscle adaptations are fundamental to many of the health benefits of exercise training, the common and sex-specific molecular regulators that mediate these adaptations remain to be fully elucidated. Methods: To this end, we leveraged skeletal muscle multi-omics data generated by the Molecular Transducers of Physical Activity Consortium (MoTrPAC), where 6 month-old male and female rats endurance trained for 1, 2, 4, or 8 weeks. Our objective was to identify shared and sex-specific multi-omic molecular responses to endurance training in skeletal muscle, and relate them to phenotypic adaptations. Results: We identified largely sexually-conserved transcriptomic and proteomic pathway enrichments in the gastrocnemius, which correlated with skeletal muscle responses from a published exercise study in humans. We uncovered sex-consistent post-translational modifications, including decreased oxidation of MYH2 and deacetylation of the β-oxidation enzyme HADHA. Pathway enrichment analyses revealed sex-specific remodeling across the acetylome, redox proteome, and phosphoproteome; females decreased mitochondrial protein cysteine oxidation and increased mitochondrial cristae proteins, indicative of enhanced redox buffering and mitochondrial efficiency. Despite decreases in cysteine oxidation of key mitochondrial proteins, females displayed increases in the cysteine oxidation of proteins involved in glucose catabolism relative to males after 8 weeks of training, suggestive of sex-biased subcellular reactive oxygen species generation. Males demonstrated earlier induction of mitochondrial transcripts and predicted activation of mTOR. Although the increase in mitochondrial protein abundance was more modest in males, there was greater oxidation of mitochondrial proteins in response to training compared to females. Conclusions: This work shows a large portion of the adaptive response to endurance training in skeletal muscle is shared between females and males, while there are distinct and nuanced sex-specific adaptations that are evident, particularly at the level of post-translational regulation.

The biology of individual lipid species and their relevance in Alzheimer’s disease (AD) remains incompletely understood. To explore the lipidomic biomarkers associated with cognition function and neuropathological changes in AD, we utilize non-targeted mass spectrometry on 316 post-mortem brains from participants in the Religious Orders Study (ROS) or Rush Memory and Aging Project (MAP) cohorts classified as control, asymptomatic AD (AAD), or symptomatic AD (SAD), and integrate the lipidomics data with untargeted proteomics from the same individuals. We find that lysophosphatidylethanolamine (LPE) and lysophosphatidylcholine (LPC) species are significantly lower in SAD than controls or AAD. Lipid-protein network analyses reveal that LPE/LPC modules are significantly associated with protein modules involved in MAPK/metabolism, post-synaptic density, and cell-ECM interaction pathways, and correlate with better antemortem cognition and reduced AD neuropathology. Particularly, LPE 22:6 [sn-1] is significantly decreased SAD and exerts a pronounced influence on protein changes relevant to neurotransmitter-driven post synaptic changes and plasticity compared to other lysophospholipids species. These findings suggest LPE 22:6 as a potential lipid signature and therapeutic target for AD.

Endurance exercise is broadly beneficial to cardiopulmonary function, with these benefits thought to be driven by extrapulmonary factors rather than direct structural changes in the lungs. Thus, to address how endurance exercise training and sex impact molecular responses in the lungs, we used a multi-omics approach to study 6-month-old Fischer 344 rats that undertook a progressive endurance treadmill training protocol for 1 to 8 weeks. Specifically, we reannotated publicly accessible transcriptomics, metabolomics, proteomics and phosphoproteomics data from the Molecular Transducers of Physical Activity Consortium and integrated newly analyzed acetylproteomics data to assess multi-omic sex differences in sedentary and treadmill trained rats. Female rats displayed enrichment in immune-related features and pathways at the transcriptome and proteome level that were largely maintained with training. However, both sexes exhibited decreases in immune pathway activity following 8 weeks of training, although the effect was more pronounced in males. Shared responses to training included increased enrichment in transcriptomic pathways related to type I alveoli, proteomic pathways related to cilia, and decreased acetylation of pathways linked to mitochondrial function. Furthermore, features known to be enriched in lung diseases were attenuated with training in both sexes. Together, our findings provide novel insight into responses to endurance exercise training in the healthy rat lung and may offer translational insight into sex-specific differences in lung disease pathogenesis and treatment.

The rapid emergence of divergent SARS-CoV-2 variants led to a 2023–2024 update of the COVID-19 mRNA vaccine to a monovalent version containing the XBB.1.5 SARS-CoV-2 spike antigen. To determine the durability and breadth of the antibody responses after immunization, we analyzed antibodies and memory B cells from 24 individuals before and after a single XBB.1.5 mRNA vaccine dose for up to 6 months. Using a live virus neutralization assay, we found that the XBB.1.5 vaccine improved the magnitude and breadth of antibody neutralizing activity against the ancestral SARS-CoV-2 strain (WA1), BA.5 strain, and XBB.1.5 Omicron variants. Durable WA1 and XBB.1.5 IgG spike protein binding antibodies were induced with an estimated half-life of 703 and 531 days (at day 120), respectively. There was a greater increase of IgG1 and IgG4 binding antibodies against the XBB.1.5 spike protein compared with the WA1 spike protein postvaccination. A high proportion of antibodies were cross-reactive against both WA1 and XBB.1.5 strains, as determined by serum depletion and memory B cell analysis, and this cross-reactivity was durable. Last, we evaluated the neutralizing activity of these antibodies against more contemporary circulating Omicron strains and observed reduced cross-reactivity to KP.2 and KP.3 compared with XBB.1.5 6 months after vaccination. These data show that the XBB.1.5 COVID-19 vaccine promotes more durable binding and neutralizing antibodies than prior ancestral WA1 or bivalent vaccines. However, divergent Omicron variants with mutations in the spike protein were able to evade these neutralizing antibodies, emphasizing the need for periodic consideration of COVID-19 vaccine reformulation.