Ut Southwestern Medical Center

NIH Research Projects · FY 2026 · 2025-07

PROJECT SUMMARY. Neurodegenerative disorders represent major sources of human suffering, yet the factors influencing disease severity remain poorly understood. Sex has been implicated as one such factor, yet there remains a considerable gap in our understanding of how sex hormones affect neurodegenerative processes. Retinitis pigmentosa (RP) is a retinal neurodegeneration in which photoreceptors undergo a progressive and irreversible degeneration leading to blindness. We recently discovered that females have a worse outcome than males in a mouse model of RP caused by the rhodopsin P23H mutation (Rho P23H), the most common cause of autosomal dominant RP in humans. Further, we showed that this association is caused by an adverse effect of circulating female sex hormones on retinal neurons, which can be ameliorated by depletion of these female hormones. RP can be caused by a wide diversity of genetic mutations, which creates a daunting obstacle to therapeutic targeting of each individual mutation. However, many of these mutations result in similar phenotypes and converge on shared downstream pathways leading to photoreceptor neurodegeneration. The objective of this proposal is to determine the molecular mechanisms by which female sex hormones adversely impact the severity of photoreceptor neurodegeneration. To investigate how the female sex hormones play a role in this common pathway of photoreceptor neurodegeneration, we propose to expand on our work in the Rho P23H RP mouse. We hypothesize that female sex hormones interact with key cell death and cell stress pathways downstream of the genetic mutation, to worsen photoreceptor degeneration. The findings from these aims will provide novel insights into how sex hormones modulate the pathogenesis and severity of neurogenerative disease. Results from these aims will identify the precise female hormones and any interconversions of these hormones driving advanced retinal degeneration in RP, and the mechanism by which this occurs. These findings have far-reaching implications for clinical trial design, such as sex stratification, hormonal medications, consideration of pre-, peri- , and post-menopausal states, and the use of hormonal therapy in females with certain neurogenerative disorders such as RP.

NIH Research Projects · FY 2025 · 2025-07

The mission of the UT Southwestern Perot Family Scholars Medical Scientist Training Program is to train the next generation of physician-scientists for a variety of medical and scientific careers that use the unique perspective of their combined MD and PhD training. The objectives are to provide integrated training leading to both MD and PhD degrees in eight years, with a high proportion of students who enter the program completing both degrees. The rationale for the program is that physician-scientists are uniquely positioned to translate laboratory discoveries to clinical practice (“bench to bedside”) and to use clinical observations to identify knowledge gaps and research opportunities (“bedside to bench”). Yet despite their widely acknowledged importance for bidirectional translation of biomedical research, clinician-scientists make up a decreasing proportion of the physician workforce. Additionally, MD-PhD graduates are needed to replenish the aging physician-scientist workforce. Program activities that promote skills development include rigorous didactic and experiential clinical and research training, emphasizing active learning in the clinic and the laboratory rather than passive learning in the classroom; extensive horizontal and vertical mentorship to foster physician-scientist identity, reinforced by abundant peer- and near-peer interactions to promote a sense of community among trainees; and robust training in scientific rigor and reproducibility as well as the responsible conduct of research. Interactions with role-model physician-scientist mentors at different career stages prepare trainees for transitions to a variety of MD-PhD careers. Twenty-four trainees out of a total of 96 training grant-eligible students will be appointed for 24 months during the first two years of medical school, with rare exceptions. Intended outcomes include timely completion of both MD and PhD degrees by a high proportion of trainees, who will continue to pursue research-related biomedical careers in academia and industry following their MD-PhD training, with continued research training during residency where applicable.

NIH Research Projects · FY 2025 · 2025-07

Down syndrome (DS) is caused by complete or partial trisomy of human chromosome 21 (Hsa21). It comprises a complex phenotype of over 80 features including metabolic dysregulation and a marked reduction in both neurogenesis and brain size. The specific molecular mechanisms by which trisomy of Hsa21 gives rise to DS pathologies remain unknown. Identifying the dominant genes or molecular pathways involved would open the possibility for developing targeted therapies to improve the lives of affected individuals. Here, we propose combining CRISPR interference (CRISPRi) technology with induced pluripotent stem cells (iPSCs) derived from individuals with DS (3S-iPSCs) to develop a selectable screen for Hsa21 genes that contribute to DS pathologies. The complexity of the DS phenotype has been a roadblock to definitive identification of genes that play prominent roles in the syndrome. Our preliminary data show evidence of two important, fundamental differences between DS cells and isogenic, euploid controls that can be detected and quantified at the single-cell level, thereby opening the door widely for the development of high through-put screens for gene identification. One of these key differences, elevated mitochondrial membrane potential (∆ψm), is evident in 3S-iPSCs in the pluripotent state; the other, reduced ability to commitment to a neuroectodermal (NE) lineage, is directly relevant to what is arguably the most prominent feature of DS, that of intellectual disability. The goal of this R21 proposal is to build the tools necessary to carry out CRISPRi-based, high throughput screens to identify genes on Hsa21 that contribute to DS pathologies at the earliest stages of development. Aim 1 will use TALEN-nuclease directed editing to integrate a transgene expressing dCAS9-KRAB into a safe-harbor locus in 3S-iPSCs. Aim 2 will focus on designing guide RNAs that achieve graded repression to more closely model a reduction in gene dosage from the trisomic three copies to the normal two copies. Successful completion of our proposed studies will provide proof-of-concept for our approach to functional gene identification in DS and lay the foundation for expanding to comprehensive and combinatorial screens of the full coding and non-coding content of Hsa21. It will also make a powerful and flexible tool available to the DS research community due to the totipotency of iPSCs.

NIH Research Projects · FY 2025 · 2025-06

Abstract Although CDK4/6 inhibitors (in combination with an endocrine therapy) are the preferred systemic therapy for treatment of patients with metastatic, Estrogen Receptor-positive (ER+) breast cancer (BC), development of resistance to CDK4/6 inhibitors is universal. Therefore, development of novel therapeutic strategies to target CDK4/6 inhibitor resistance remains an unmet clinical needs. To probe the dependency of ER+ BC cells and tumors on CDK4 and 6 for growth, we profiled a panel of ER+ BC cell lines, patient-derived xenografts (PDXs) and primary tumors from ER+ BC patients for expression of these proteins. We observed near universal expression of CDK4 but no detectable expression of CDK6 in ER+ BC samples. However, overexpression of CDK6 (and not CDK4) is a common mechanism of resistance to CDK4/6 inhibitors in ER+ BC patients. Using a panel of ER+ BC cell lines engineered to overexpress CDK6, we showed that CDK6 overexpression confers resistance to palbociclib although palbociclib completely blocked phosphorylation of pRB in these cells. Furthermore, overexpression of a kinase-dead (K43M) mutant of CDK6 (referred to as CDK6-KD) was sufficient to confer palbociclib resistance both in vitro and in vivo. Our mechanistic investigations revealed that CDK6 overexpression promotes transcriptional reprogramming of BC cells in a kinase-independent fashion and that the non-kinase function of CDK6 is a key driver of palbociclib resistance. Finally, we showed that CDK4/6-D, a CDK4/6-selective proteolysis targeting chimera (PROTAC), which causes targeted degradation of CDK6 (and CDK4), and eliminates both kinase and non-kinase functions of CDK6, exerts potent tumor growth inhibition in CDK6 high, palbociclib-resistant ER+ BC xenograft models. Supported by our preliminary data, we hypothesize that high CDK6 in ER+ BC cells causes CDK4/6 inhibitor resistance, at least in part, through kinase-independent transcriptional reprogramming, and that pharmacological targeting of both the kinase and non-kinase of CDK6 is necessary to inhibit the growth of these tumors. Specific Aim 1: To define the roles of kinase and non-kinase functions of CDK6 in conferring palbociclib resistance in a panel of CDK6-high, palbociclib-resistant cell line- and patient-derived xenograft (PDX) models of ER+ BC using advanced 3D genomic and machine learning approaches. Specific aim 2: To evaluate the in vivo efficacy of a CDK4/6-selective degrader (CDK4/6-D) in a panel of CDK6- high, palbociclib-resistant ER+ BC xenografts, both as a single agent and in combination with endocrine therapy. Specific Aim 3: To evaluate the efficacy of CDK4/6-D in a panel of CDK6-high, palbociclib-resistant PDXs engrafted in a unique NeoThy humanized mouse model, both as a single agent and in combination with endocrine therapy and checkpoint inhibition. Our novel therapeutic approach has the potential to improve clinical outcomes in many BC patients.

NIH Research Projects · FY 2025 · 2025-06

This new application is to establish the North Texas Alzheimer’s Disease Research Center (NT-ADRC) which will serve North Texas, one of the most-populous catchment areas in the country. This application describes how we plan to build a unique and interconnected multidisciplinary center that will create an environment that enables and supports innovative research, training and eventually enhance the care and lives of those dealing with Alzheimer's Disease (AD) and related dementias (ADRD). It is built on a strong foundation of AD and ADRD research and will synergize and connect multiple AD/ADRD investigators and their programs at University of Texas (UT) Southwestern Medical Center, UT Dallas, and UT Arlington. The NT-ADRC will benefit from the strong institutional support to advance AD and ADRD research, our long history of studying cardiometabolic factors, particularly hypertension, in AD/ADRD, our recent expansion of the cutting-edge neuroimaging, informatics, machine learning, and artificial intelligence (ML/AI) capabilities in neurosciences, and the recruitment of multiple key leaders in the Neurocognitive space. The NT-ADRC thematic focus is on examining the roles and mechanisms of hypertension and other cardiometabolic factors in the early phases of AD/ADRD, developing innovative biomarkers for early detection, and identifying related therapeutic targets. We have an exceptional engagement in our diverse communities that is amplified by the participation of Parkland Health and its community ties. We also create novel ways to enhance inclusiveness in ADRD research particularly Lumbar Punctures acceptance and Brain Donation enrollment. These amplify the Center’s abilities to create a highly diverse UDS cohort representative of our diverse population as evidenced by our affiliated studies (Dallas Heart Study and the NIA-funded Post-COVID Neuro-Cognitive Manifestations in Older African Americans). The NT-ADRC Center addresses multiple elements in the National Alzheimer's Project Act (NAPA) and its Implementation Milestones by enabling ongoing and new research in the preclinical and prodromal stages centered around the discovery of novel disease mechanisms underlying the role of hypertension and other cardiometabolic factors in increasing the risk of AD/ADRD; implementing novel ML/AI approaches to biomarker developments; and paving the way for novel therapeutic developments. The NT- ADRC will enhance scientific and clinical collaborations by sharing of biosamples and data with local and national investigators, dissemination of research findings to professional and lay audiences and create education opportunities for researchers, clinicians, learners, and the general North Texas community focused on enhancing caregivers’ education and support. Our commitment to diversity, innovation and rigor to ADRD research will build a new local environment of cutting-edge research and training. It will also position us to make important contributions to the ADRC network and to advance the national agenda towards meeting the NAPA goals.

NIH Research Projects · FY 2026 · 2025-06

PROJECT SUMMARY This Mentored Research Scientist Career Development Award proposal is designed to provide advanced training, expert mentoring, and hands-on research experience that will facilitate my successful transition to research independence. My primary career goal is to establish myself as a translational neuroscientist with specific expertise in the application of in vivo Positron Emission Tomography (PET) to animal models of stress and aging. To achieve this goal, I propose a comprehensive five-year plan designed to provide rigorous training in four key areas: 1) Basic training in the biology of stress and aging; 2) Practical training in molecular, behavioral, and synaptic assessments, including training in PET methodology; 3) Advanced biostatistical training; 4) Career development and mentorship to facilitate the transition to research independance. Integral to this training plan is completion of a novel research project which uses state-of-the-art in vivo PET imaging techniques that allow for the longitudinal evaluation of synapses in an animal model of chronic stress and aging. Using clinical neuroimaging in animal models provides an important translational bridge between basic science and the clinic, facilitating the dissemination of findings from bench-to-bedside. Typical aging is associated with widespread brain changes that can contribute to cognitive decline. Stress can similarly impact brain physiology, and it is hypothesized that these stress- and age-related brain alterations are driven in part by synaptic changes, including synaptic loss. Further, evidence suggests chronic stress may accelerate brain aging and contribute to premature cognitive decline. Previously, the hypothesis that stress accelerates synaptic aging could not be directly tested in vivo because no method existed for measuring synapses in the living human brain. However, this is no longer the case: with the recent development of radiotracers targeting synaptic vesicle protein 2A (SV2A), PET quantification of tracer binding provides a method for estimating synaptic density in vivo. Here, we propose using SV2A PET imaging in an animal model. Specifically, we aim to determine the effect of chronic stress on synaptic density using in vivo SV2A PET (Aim 1), investigate the impact of age on stress-induced synaptic changes (Aim 2), and explore relationships between stress- and age-related synaptic changes with cognitive function (Aim 3). Results of this study will provide potentially critical insight into synaptic mechanisms contributing to stress-accelerated brain aging and relationships with functional and cognitive decline. Completion of the proposed training and research plan will optimally prepare me for a career as an independent neuroscientist and translational neuroimager, and ultimately establish a research program capable of improving our understanding of the biological underpinnings of stress-related neuropathology, aiding the development of novel treatments, and reducing stigma and suffering.

NIH Research Projects · FY 2025 · 2025-06

PROJECT SUMMARY Brain metastasis is a complex process which involves the tumor cells, resident cells, and infiltrating immune cells. Glial cells are long lived and have been shown to react to both radiation therapy and peripheral stimuli such as systemic inflammation. Microglia can become continually reactive, inciting brain damage or creating a prolonged inflammatory environment. As radiation therapy is oftentimes the first treatment option for patients with brain metastasis, how glial cells and the broader niche responds, is critical to study. Various functions of microglia including their ability to engage in complement signaling is fundamental to their proper functioning. How radiation influences this pathway is not well understood. Furthermore, many people today have systemic inflammation, which will also influence glial responsiveness to treatment and further cancer growth. An emerging body of literature has demonstrated that non-adaptive immune cells have a form of ‘memory’ through epigenetic regulation, which results in either an increased or decreased ability to react to subsequent stimuli. How the systemic effect of inflammation, modeled by beta glucan, influences radiotherapy outcomes is not fully understood. In the brain metastasis context, many peripheral immune cells will migrate to the brain. Ccr2 positive myeloid cells increase proportionally in mice treated with radiation when compared to control mice. To gain a better understanding of what is occurring within the treated brain metastatic niche this proposal seeks to examine 1) the mechanisms through which radiation therapy-stimulated complement signaling in microglia and infiltration of peripheral myeloid cells regulate brain metastasis progression and 2) discover how systemic inflammation influences radiotherapy outcomes and how systemic inflammation could be treated to ensure proper microglial functioning. Utilizing cellular imaging, qPCR, and ATAC- and RNA-seq we will explore how microglial cells are trained by radiation, systemic inflammation, and how this could be treated. To discover how proper functioning of microglia is critical to positive outcomes we will use genetic knockouts. The overarching hypothesis is that radiation alone and radiation within the context of systemic inflammation leads to alterations within microglia which contribute to a pro-inflammatory and overactive phenotype. To test this hypothesis two specific aims will be explored. Experiments under Aim 1 will 1) if complement signaling is a factor in brain metastasis outgrowth and 2) uncover the role of Ccr2+ infiltrating myeloid cells within the brain metastatic niche. Aim 2 will focus on 1) how systemic inflammation trains microglia, 2) if the mechanism behind training is PI3K-AKT, and 3) if we can treat systemic inflammation trained microglia with drugs that work synergistically with radiotherapy. Collectively these data will reveal mechanisms important to metastatic growth within mice exposed to radiation alone or radiation in concert with systemic inflammation, focusing primarily on microglia. This research will have broad implications for understanding what is occurring within the brain metastatic niche as well as for potential avenues for therapeutic potential.

NIH Research Projects · FY 2025 · 2025-06

eDyNAmiC (extrachromosomal DNA in Cancer) Human genes are arranged on 23 pairs of chromosomes, but in cancer, tumour-promoting genes can free themselves from chromosomes and relocate to circular, extrachromosomal pieces of DNA (ecDNA). These ecDNA do not follow the normal “rules” of chromosomal inheritance, enabling tumours to achieve far higher levels of cancer-causing oncogenes than would otherwise be possible, and licensing cancers with a way to evolve and change their genomes to evade treatments at rates that would be unthinkable for human cells. The altered circular architecture of ecDNAs also changes the way that the cancer-causing genes are regulated and expressed, further contributing to aggressive tumour growth. These unique features make ecDNA-containing cancers especially aggressive and difficult to treat. Cancer patients whose tumours harbour ecDNA have markedly shorter survival. Despite being first seen over fifty years ago, the critical importance of ecDNA has only recently come to light, and the scale of the problem is substantial. ecDNAs are present in nearly half of all human cancer types and potentially up-to a third of all cancer patients. The collective current understanding of how ecDNA form, how they function, how they move around the cell, how they evolve to resist treatment, how they impact the immune system, and how they can be effectively targeted are lacking. We bring together an internationally recognized, pioneering interdisciplinary team of cancer biologists, geneticists, computer scientists, evolutionary biologists, mathematicians, clinicians, and patient advocates to boldly create novel insights and resources and to provide transformative solutions to one of Cancer’s Grand Challenges. A core team of experienced and productive ecDNA investigators will work with new investigators in the ecDNA and cancer fields to bring completely new perspectives and approaches to this daunting challenge. By bridging cutting-edge and diverse approaches and insights from cancer genomics, yeast genetics, epigenomics, artificial genome synthesis, longitudinal patient tracking, combinatorial and machine learning algorithms, mathematical modelling, immunobiology, and innovative chemistry we will develop a new understanding of the role of ecDNA in cancer, and we will find new ways to drug the undruggable. This bold programme, which consists of 7 work packages and a committed international infrastructure, generates new and unusual collaborations that would simply be impossible under any other type of funding mechanism. Our programme endeavours to foster bold innovative solutions to one of the hardest problems in cancer and to one of the greatest challenges facing cancer patients.

NIH Research Projects · FY 2026 · 2025-05

PROJECT SUMMARY Gametogenesis, the intricate process of generating cells specialized in sexual reproduction through meiosis, necessitates a profound alteration in gene expression, organelle homeostasis, and programmed cell development and differentiation in the context of targeted proteolysis. Autophagy—a highly-conserved lysosomal degradation process—plays a crucial role in cell survival under diverse stress conditions and has emerged as a pivotal player in governing reproductive and developmental stages, including gametogenesis. In the yeast Saccharomyces cerevisiae, autophagy is active and essential at multiple stages including meiosis entry, DNA replication, meiosis exit and the subsequent daughter cell membrane biogenesis. Phenotypes associated with failed meiosis exit include aberrant spindle pole body (SPB, yeast centrosome) formation and chromosome segregation. To date, Cdc14, a conserved cell cycle phosphatase is the only meiosis-specific autophagy regulator identified. Cdc14 is involved in guiding autophagy at anaphase I to remove Rim4 selectively, a meiosis-specific RNA binding protein (RBP) that inhibits translation. This finding, from my laboratory, reveals a novel link between autophagy and temporal meiotic translation at a specific stage of meiosis, meiosis II, while critical autophagy functions and regulations at other stages of gametogenesis are yet to be revealed. On the other hand, yeast gametogenesis in nature is exposed to stresses, such as starvation, heat, and oxidation. I recently discovered meiosis-specific stress granules (SGs) triggered by stressors, including heat and oxidation. The SGs typically sequester mRNAs to inhibit translation until being disassembled during recovery, and autophagy degrades the persistent SGs. Intriguingly, meiotic SGs discovered in our study exhibit features including high heat sensitivity and autophagy resistance. Moreover, the capacity of meiotic SG formation correlates with the efficiency of meiotic DNA replication and gametogenesis. Nevertheless, the functions and regulation of meiotic SGs remain mysterious. Employing a multidisciplinary approach encompassing genetics, biochemistry, cellular imaging, proteomics, and computational methods, this proposal will investigate three overarching directions: D1) Determine how autophagy promotes gametogenesis; D2) Determine how gametogenesis regulates autophagy; and D3) Investigate how stresses affect autophagy and gametogenesis through stress granules (SGs). My investigation will primarily focus on the steps of meiosis but also cover pre-meiotic and post-meiotic stages of yeast gametogenesis. This research is poised to yield interdisciplinary conceptual breakthroughs in autophagy, gametogenesis, and stress responses. The revealed mechanisms, once confirmed in higher eukaryotes, including humans, will shed light on infertility, miscarriage, and specific human disorders like Turner syndrome (monosomy X, frequency: 1/2,500 newborn girls) and Down syndrome (trisomy 21, frequency: 1/800 newborns).

NIH Research Projects · FY 2026 · 2025-05

PROJECT SUMMARY/ABSTRACT The need for new antimicrobials is increasingly urgent. The rate of multidrug resistant pathogens continues to increase, leading to significant morbidity and mortality throughout the world. Furthermore, the current pipeline for new antimicrobials remains very narrow. The CDC has identified a number of Gram-negative pathogens as Urgent or Serious threats. A new paradigm in antibiotic discovery and design has recently been shown effective against numerous bacteria. This new approach is based on a platform technology called peptide- phosphorodiamidate morpholino oligomers (PPMOs). PPMOs are synthetic DNA mimics that bind to RNA in a sequence-specific, antisense manner and inhibit translation of target bacterial genes. PPMOs have already been used successfully to kill a variety of bacterial pathogens including the Gram-negative bacteria Pseudomonas aeruginosa, Salmonella typhimurium, Burkholderia cepacia complex (Bcc), Acinetobacter baumannii, Klebsiella pneumoniae and Escherichia coli. PPMOS are bactericidal in culture, and reduce bacteremia and improve survival in animal models of infection. In addition, PPMOs retain activity in both multidrug-resistant strains and biofilm settings. The mechanisms of resistance to PPMOs remains understudied. In addition, it is not clear whether common themes of resistance will be seen across many of these medically important pathogens. The goal of this project is to determine the mechanism of resistance for lead PPMOs in the pathogens Klebsiella pneumoniae, Acinetobacter baumannii and members of the Bcc. These representative members of different Proteobacteria classes will allow us to broadly study possible shared mechanisms of resistance. Utilizing species-specific lead PPMOs that target the same essential gene, mutants will be generated and characterized. Resistance circumventing strategies will be developed depending on the mechanism of resistance. The hypothesis is that drivers of resistance will be related to peptide- mediated entry or target gene sequence mutations and strategies to overcome these will be tested through new PPMO design and testing.

NIH Research Projects · FY 2026 · 2025-05

PROJECT SUMMARY/ABSTRACT The ability to precisely track gene expression is fundamental to unraveling the complexities of brain function, elucidating the mechanisms underpinning neurological disorders, and devising targeted diagnostic and therapeutic interventions. While bioluminescent imaging (BLI) has traditionally been the primary method for monitoring gene expression in living animal models, it has inherent limitations such as limited optical signal penetration and reliance on external detectors. In contrast, magnetic resonance imaging (MRI) offers exceptional capabilities for deep tissue imaging and full-brain coverage. However, existing molecular MRI-based methods have been hampered by low signal-to-noise ratios or potential cell toxicity when sensor overexpressed. In this proposed project, we introduce an innovative optical MRI (oMRI) technique for the time-resolved visualization of gene expression patterns within living brains. This novel imaging approach harnesses the strengths of bioluminescent imaging for molecular specificity and sensitivity, in conjunction with MRI's capacity for 3- dimensional imaging and high spatiotemporal resolution. We achieve this by directly associating bioluminescent signals with hemodynamic MRI signals using optogenetic tools, allowing us to map the expression of target genes across both space and time. In Aim 1, we will implement the oMRI strategy to image rhythmic clock gene expression in living mouse brains, building on our already validated luciferase-opsin pair design. We expect the oMRI technique to detect differential Per2 expression levels in mice brains exposed to 12-hour light and dark cycles. This pioneering imaging strategy will establish a foundation for tracking Per2 and other core clock genes in the brain and significantly advance Alzheimer's disease research. In Aim 2, we will advance the oMRI toolkit and develop dual-channel gene expression imaging. This will allow us to study the longitudinal interactions of multiple molecular events during AD progression, particularly the relationship between clock gene disruptions and other AD pathological events. We anticipate that this novel MRI technique will revolutionize our understanding of the circadian system's role in AD development, paving the way for discovering biomarkers for early AD diagnosis and intervention.

NIH Research Projects · FY 2025 · 2025-05

PROJECT SUMMARY For reasons still not well understood, women have a higher prevalence of knee articular cartilage degeneration than men specially after menopause. Sex-specific gait mechanics, muscle strength, metabolism, and sex hormones are key contributors to this health disparity. However, despite growing efforts to reveal sex differences in cartilage, the influence of both the biological sex and sex hormones in modulating cartilage anabolism and catabolism are still unclear, especially during age-related hormonal changes (contraceptive use, menopause, and hormonal replacement therapy in women). As cartilage structure results from chondrocyte mechanotransduction, we propose that the key to sexual dimorphism in cartilage and the exacerbation of risks for catabolism after menopause in women are related to how chondrocytes respond to mechanical stimulation depending on both the biological sex and sex hormones. Defining how biological sex and sex hormones modulate chondrocyte mechanotransduction will help identify new targets to treat and prevent age-related cartilage degeneration, especially in women undergoing hormonal changes. Moreover, answering these questions will be foundational in understanding how sex hormones affect cartilage in gender minorities, specifically the transgender population going through gender-affirming intervention, for whom changes in musculoskeletal health after transition are understudied. The calcium-permeant mechanosensitive channels PIEZO1 and TRPV4 are well-known mediators of chondrocyte mechanotransduction and involved in cartilage anabolism and catabolism. According to our supporting data, their gene expression in response to physiological levels of cyclic compression are influenced by the biological sex and 17-β Estradiol (E2), and E2 decreases the calcium influx induced by the pharmacological activation of TRPV4 and PIEZO1. Therefore, we hypothesize that E2 affects chondrocyte mechanotransduction by regulating the activity and downstream effect of PIEZO1 and TRPV4 in a sex-specific manner. Since we previously found sex differences in the extracellular matrix (ECM) of adult bovine cartilage (equivalent to human premenopausal ages), we will investigate sex differences in chondrocyte mechanotransduction without the confounding effect of the ECM and at premenopausal ages. Here, we will isolate chondrocytes from healthy human cartilage from postmortem donors (20-40 years old) to grow them in 2% agarose discs and test our hypothesis by defining the sex-specific and E2-regulated role on the activation and downstream effect of PIEZO1 and TRPV4 (Aim 1) and defining the effect of biological sex and E2 in chondrocyte mechanotransduction mediated by PIEZO1 and TRPV4 (Aim 2). In Aim 1, we will (1.1) measure calcium intake transients in absence or presence of a premenopausal concentration of E2, and (1.2) perform phosphoproteomic and bulk-RNAseq analysis of chondrocytes treated with specific PIEZO1 and TRPV4 agonists (Yoda-1 and GSK101, respectively) in the absence or presence of E2. In Aim 2, we will stimulate chondrocytes with cyclic compression at physiological levels (8-10% strain) to activate TRPV4 (2.1) or a hyper physiological level (70% strain) to activate PIEZO1 (2.2) in absence or presence of E2. The most significant differentially expressed genes modulated by E2 after activation of each channel in Aim1 will be transiently silenced with siRNA before applying mechanical stimulation. We will assess the expression of selected anabolic and catabolic markers. We propose that testing our hypothesis first in premenopausal aged samples is key to later expand to postmenopausal samples. Data obtained in this E2-focused study will be foundational for a future multi-hormonal study.

NIH Research Projects · FY 2026 · 2025-05

PROJECT SUMMARY. Retinal degenerative diseases are characterized by the loss of the photoreceptors, the cells within the eye that convert light into signals for vision. Photoreceptors in the mammalian eye are unable to divide or regenerate, and once they are damaged, they cannot be replaced. Thus, patients with retinal degenerative diseases become blind over time and undergo a substantial decrease in their quality of life, as they are no longer able to read, drive or perform independent daily activities. For most of these diseases, treatment to delay photoreceptor cell death and preserve visual response is unavailable. Cellular pathways that promote photoreceptor identity and / or survival have important implications for future clinical therapeutics, such as cellular transplants and reprogramming. Gene activity is controlled by transcriptional regulators, and one group of transcriptional regulators, the PRDM family, has been shown to drive and maintain cell state transitions in a wide range of tissues and cell types. Of interest, one member of the PRDM family, PRDM13, is associated with retinal abnormalities, but no mammalian models existed to examine its activity on the formation of proper retinal cell identities. The overall goal of our research is to determine how PRDM13 regulates retinogenesis and its association with North Carolina Macular Dystrophy. Our central hypothesis is that PRDM13 acts during a critical window of retinal development to cause a shift in cell fate determination disrupting formation of photoreceptors. Completion of the proposed aims will further our understanding of mammalian retinal development by establishing mechanisms for cell fate decisions guided by PRDM13 transcriptional regulation, and how these contribute to healthy retinogenesis.

NIH Research Projects · FY 2026 · 2025-05

PROJECT SUMMARY/ABSTRACT Obesity is a major risk factor for hypertensive disorders of pregnancy (HDP). The underlying mechanisms are largely unclear, but maternal vascular endothelial dysfunction is likely involved. Endothelial dysfunction in HDP could be attributed to 1) alterations in the L-arginine/nitric oxide (NO) pathway, and 2) an increase in endothelin-1 (ET-1). Additionally, augmented sympathetic vasoconstriction may also contribute to HDP. Chronic (repeated) whole-body heat exposure has been shown to increase NO bioavailability, decrease ET-1, and cause functional and structural adaptations in the vasculature. All these can improve vascular function, attenuate sympathetic (re)activity, lower blood pressure (BP), and reduce cardiovascular risk in non-pregnant women. Whether this is also true after regional (lower leg) heating in high-risk pregnant women is unknown. Our central hypothesis is that chronic lower leg heating will be effective in improving vascular endothelial function and attenuating sympathetic vasoconstriction, leading to a reduction of the risk for HDP in pregnant women with obesity. The overarching goal of this proposal is to determine the vascular and neural effects of chronic lower leg heating during pregnancy in women with obesity. We plan to enroll 118 pregnant women with obesity between 12-14 weeks of gestation and randomly assign them to either an intervention group or a control group (1:1 ratio). Participants in the intervention group will perform 16 weeks of home-based lower leg heating via water immersion up to the knee in a circulated bath (water 42°C, 4 times/week, 45 min/session), whereas those in the control group will immerse their legs in a thermoneutral water bath (33°C) at the same frequency and duration. Participants will be evaluated at baseline and then at 28-30 weeks of gestation. Aim 1 will determine the effects of chronic lower leg heating on maternal vascular function and surrogate markers of HDP. We will measure brachial artery flow-mediated dilation and placental perfusion (ultrasound). Blood samples will be collected for the estimation of NO (via nitrate and nitrite analysis), L-arginine, asymmetric dimethylarginine, 3-nitrotyrosine, and ET-1. Angiogenic factors, in combination with ambulatory BP, will be used as surrogate markers of HDP. Aim 2 will determine the effects of chronic lower leg heating on sympathetic vasoconstriction and BP. We will measure sympathetic activity (microneurography), leg vascular conductance, and neurovascular transduction. Sympathetic reactivity will be assessed during a graded upright tilt. We will also measure 24-h ambulatory BP. The relationship between sympathetic (re)activity and BP will be evaluated. Finally, we will determine the association between changes in sympathetic vasoconstriction and endothelial function after intervention. Findings from this project will provide insight on the extent and potential mechanisms of how chronic lower leg heating works for improving vascular endothelial function and sympathetic vasoconstriction in pregnant women with obesity. Results obtained will set a foundation for future large multicenter clinical trials to determine the efficacy and generalizability of home-based lower leg heat therapy as a safe, ease-of-use, cost-effective, and non-drug approach for reducing the risk of HDP.

NIH Research Projects · FY 2025 · 2025-05

Modified Project Summary/Abstract Section The podocytopathies are a group of glomerular diseases that affect the kidney’s ability to filter the blood and often lead to kidney failure. Healthy podocytes cover the glomerular capillaries with thousands of extensions called foot processes that interdigitate with one another and maintain their elaborate cell shape by tightly regulating their actin cytoskeleton. Podocytes respond to insults in a typical fashion by undergoing foot process effacement, a dramatic shift in podocyte morphology and the disappearance of the intricate foot processes, which often associates with the “actin mat”, an actin condensation at the bottom of the effaced areas. We recently used super-resolution imaging to study the podocyte actin cytoskeleton in 3D in both healthy and diseased conditions. We showed that healthy podocyte foot processes contain non-contractile actin cables, while contractile cables are maintained high in the cell bodies. In contrast, injured podocytes appear to have contractile actin cables in effacement areas juxtaposed to the glomerular basement membrane (GBM), indicating a shift in the spatial distribution of actin cables after injury. The overall goal of this proposal is to define the molecular mechanisms that regulate the various types of actin cables in podocytes and the nature of the changes that cause the contractile actin cables in the cell body to shift towards the effaced areas adjacent to the GBM after injury. In Aim 1, we will investigate the roles of the two isoactins, beta and gamma actin, in podocyte pathobiology. Podocytes express high levels of these almost-identical evolutionally-conserved isoactins. While beta actin in non-muscle cells is considered the main isoactin, as evident from the embryonic lethality when inactivated, the role of gamma actin is still elusive. we will use various kidney disease mouse models, including the gamma-actin knockout mouse, to answer some fundamental questions about the role of the two isoactins in podocyte biology. Furthermore, we will utilize a novel technique to study primary podocytes as they spread out of isolated kidney glomeruli onto a substrate-micropatterned hydrogel. This approach will allow us to study the dynamic changes in the actin cytoskeleton in injured podocytes and will shed more light on the fate of the actin mats in effaced podocytes. It will help us identifying the role of Rho small GTPases and its downstream effectors, formins, in the actin mat formation. In Aim 2, we will study the tropomyosin isoform composition in podocytes and their roles in specifying the spatial distribution of different types of actin cables in the kidney podocytes. We hypothesize that changes in tropomyosin composition in injured podocytes causes the ectopic appearance of contractile actin cables in the effaced areas, and this, in turn, is regulated by different formins. Understanding how tropomyosins regulate the various types of actin cables could provide the missing link to podocyte foot process effacement. Our goal is to expand our understanding of the molecular mechanisms that regulate the composition and dynamics of the actin cytoskeleton, a step that will help us in designing novel therapeutic approaches to directly impact podocyte foot process architecture and help cure kidney glomerular diseases.

NIH Research Projects · FY 2026 · 2025-05

PROJECT SUMMARY/ABSTRACT The interstitial lung diseases (ILD) are a group of incurable fibrotic and inflammatory disorders that confer substantial mortality risk through progressive loss of lung function. The ILDs are categorized into discrete diagnostic subtypes to inform clinical management. However, diagnostic categorization is imprecise and is unable to reliably identify individual patients at high risk for rapid progression or poor survival. Leukocyte telomere length is an emerging prognostic biomarker that consistently informs individual progression and mortality risk. In addition, a recently discovered pharmacogenomic interaction between short leukocyte telomere length and immunosuppressant medications suggests that leukocyte telomere length is also a predictive biomarker that can aid in ILD treatment selection. This proposal seeks to leverage a multicenter cohort of >4000 ILD patients to outline relevant information gained from clinical leukocyte telomere length measurement across the spectrum of fibrotic ILDs. In Aim 1, we will quantify the age-adjusted leukocyte telomere length threshold that best informs differential progression and mortality risk. Using these thresholds, we will then calculate performance characteristics to provide clinicians with discriminatory power of leukocyte telomere length in forecasting outcome risk to inform clinical decision making. In Aim 2, we will study treatment naïve fibrotic ILD patients to quantify the effect of age-adjusted leukocyte telomere length on incident progression and near-term mortality while accounting for ILD treatment effects. This aim will quantify the independent impact leukocyte telomere length and treatment selection on ILD outcome risks. In Aim 3, we will create a risk prediction model anchored on leukocyte telomere length that informs both near-term progression and mortality risk. We will then incorporate subsequent treatment exposure to calculate the change in risk profile conferred by specific treatment selection. Successful completion of the proposed aims will create a tractable predictive tool integrating a blood biopsy with clinic-radiologic features that informs personalized ILD management without constraints of the current, imprecise ILD diagnostic categorization. In addition, results from these studies will form the foundation for future prospective cohort studies and stratified clinical trials to validate the ability of leukocyte telomere length to function as a prognostic and predictive biomarker and usher in the era of precision medicine in fibrotic ILD.

NIH Research Projects · FY 2026 · 2025-05

PROJECT SUMMARY/ABSTRACT Inborn errors of metabolism (IEMs) are genetic diseases that often cause severe illness and death in infancy or early childhood. IEMs represent the largest subset of genetic diseases in children, numbering over 1000, with a combined incidence greater than 1 in 1000. For some IEMs, early identification and intervention permit normal development, but in many cases, an incomplete understanding of IEM pathophysiology limits therapeutic options. Defects in lipoic acid (LA) metabolism represent a recently described category of IEM that causes complex metabolic disease, liver injury, and neurodevelopmental delay. Little is known about the mechanistic drivers of pathology in patients with these defects, and identification of key accumulated or deficient metabolites in vivo holds promise to inform therapeutic intervention. Lipoylation, or the addition of an LA moiety to a target protein, is required for the function of specific metabolic enzymes referred to as 2-ketoacid dehydrogenases. These enzymes are critical contributors to central carbon metabolism and include pyruvate dehydrogenase (PDH) and α-ketoglutarate dehydrogenase (α-KGDH). Lipoyltransferase-1 (LIPT1) catalyzes the transfer of an LA moiety to the 2-ketoacid dehydrogenases. LIPT1 deficiency in patients results in severe illness with restricted growth and neurodevelopmental delay. In healthy individuals, metabolic flexibility in the liver permits mobilization of energy stores during growth and development to maintain homeostasis. Hospitalizations and deaths in patients with IEMs often occur during periods of homeostatic decompensation. I hypothesize that impaired central carbon metabolism in LIPT1 deficiency restricts hepatic metabolic flexibility, destabilizing maintenance of homeostasis. Preliminary data reveal that knockout of Lipt1 in mouse livers results in acute weight loss and death 6-8 weeks after induction, while mice with developmental loss of Lipt1 in the livers display restricted growth with ~40% lethality by 20 weeks of age. With these novel, complementary models, this proposal will dissect the impact of developmental loss of lipoylation from the role of lipoylation in healthy liver physiology. If successful, it will define the metabolic alterations driven by lipoylation loss and attenuate these alterations with targeted dietary intervention to bypass blockades at both PDH and α-KGDH.

NIH Research Projects · FY 2026 · 2025-05

Project Summary/Abstract Osteonecrosis (ON) is one of the most debilitating bone diseases, with approximately 20,000-30,000 new cases diagnosed annually in the US and over 20 million worldwide. The disease progresses in 80% of patients, leading to bone collapse and osteoarthritis. Current ON treatments remain suboptimal. Non-operative treatments fail to reverse ON progression due to insufficient local bioavailability caused by poor circulation in ON-affected bone. Operative treatments aim to “Demolish and Rebuild” by removing as much necrotic bone as possible and filling the large bone void with bone substitutes, which does not improve the prognosis and raises concerns of iatrogenic bone collapse, heterotopic ossification, and pulmonary embolism. Due to treatment failures, ON patients often require total joint replacement, which is not ideal in young adults given their high physical demand and longevity. To solve the challenge, we propose to develop a novel pro-Angiogenic Bone Coating hydrogel (ABC hydrogel), which is a copolymer of gelatin and hyaluronic acid, which can provide high coupling and adjustable degrees of bisphosphonate (polyBP) and pro-angiogenic QK peptide (polyQK, VEGF mimicking peptide). Our preliminary studies found: 1) Injecting ABC hydrogel via small intraosseous needles produced a broad distribution within the necrotic bone without leakage; 2) After injection, ABC hydrogel can quickly transition to liquid at body temperature without blocking the marrow space, which is critical for angiogenesis, new repair tissue ingrowth, and avoiding pulmonary embolism; 3) The bifunctional ABC molecule can attach to the necrotic bone surface via polyBP component to inhibit the osteoclast activity and accelerate revascularization via polyQK component; 4) Although there is concern about delayed bone remodeling related to the bisphosphonate (BP) component, our preliminary results found that the ABC hydrogel treatment in ON rats effectively prevented bone deformity and allowed bone remodeling. These promising characteristics make the ABC hydrogel an excellent candidate for treating ON. This study proposes three specific aims: Aim 1 is to determine the physical, chemical, and cytocompatibility of the ABC hydrogel for ON treatment. This includes characterizations of the chemical structure and sol-gel transition, tests of the injection pressure and hydrogel distribution, and assessments of the degradation and cytotoxicity. Aim 2 is to optimize the pro-angiogenic capability of the ABC hydrogel. ABC hydrogel formulations with different QK and BP components will be studied, including evaluations of in vitro angiogenic bioactivities, ex vivo release of pro-angiogenic QK, and in vivo revascularization. Aim 3 is to optimize the antiresorption capability of ABC hydrogel to prevent early bone resorption while allowing restoration of bone remolding. ABC hydrogel formulations with maximum QK components and different BP components will be studied, including evaluations of in vitro anti-resorption properties, in vivo BP release, and in vivo ON therapeutic effects. Successful completion of this project will develop a novel bioactive hydrogel system, establish a promising cell-free strategy, and open a new direction of using a minimally invasive manner for ON treatment.

NIH Research Projects · FY 2026 · 2025-05

PROJECT SUMMARY Elevated plasma LDL cholesterol is a potent risk factor for atherosclerosis and cardiovascular complications. LDL is cleared from circulation by LDL receptors (LDLR). Importantly, the balance between internalization, recycling, and intracellular degradation of LDLR is critical in determining surface expression of LDLR, LDL clearance, and plasma LDL concentrations. A pivotal regulator of endosomal recycling of LDLR is the trimeric protein complex known as Retriever. Comprised of VPS35L, VPS26C, and VPS29, Retriever is essential for identifying cargo proteins in the endosomal compartment to initiate their recycling to the plasma membrane (PM). Over the past year, our multi-PI group leading this application has published the first high-resolution structural model of Retriever using cryogenic electron microscopy (cryo-EM), revealing unique features of this complex. Recycling of LDLR and other cargoes requires Retriever binding with several crucial ligands, including sorting nexin SNX17, the COMMD/CCDC22/CCDC93 (CCC) complex, and the WASH complex. Mutations in Retriever, CCC and WASH complex subunits are associated with hypercholesterolemia in humans. However, the molecular mechanisms linking Retriever to CCC and WASH complexes remain incompletely understood, representing a major knowledge gap in the field, hindering our mechanistic understanding of how Retriever regulates cholesterol homeostasis. Our hypothesis is that by using structural insights to precisely uncouple Retriever from these critical ligands, we can achieve a comprehensive understanding of their functional contributions to endosomal recycling and cholesterol homeostasis. Our team will combine expertise in biochemistry, structural biology, cell biology, and animal models to obtain a detailed molecular understanding of Retriever function through 3 aims: (1) Structural and biochemical analysis of Retriever binding with key ligands, (2) Functional characterization of Retriever-ligand interactions in cargo recycling using cell models, and, (3) Studies on the physiological significance of Retriever-ligand interactions in cholesterol homeostasis using in vivo models of Retriever deficiency. Altogether, completion of this work will greatly advance our mechanistic understanding of Retriever, a critical regulator of endosomal protein recycling. Furthermore, by taking advantage of our unique structural, biochemical, cellular, and animal model approaches, we will identify molecular pathways regulated by Retriever-dependent recycling that play meaningful roles in determining circulating LDL levels. These studies are of high relevance to cardiovascular health and the mission of NHLBI.

NIH Research Projects · FY 2026 · 2025-05

Background: There is broad variation in nephron endowment in humans; infants born with fewer nephrons have increased risk of chronic kidney disease (CKD) in adulthood. Nicotinamide adenine dinucleotide (NAD+) is a critical co-factor in most energy producing reactions. It is synthesized from nutritional precursors. Published work proposes that NAD+ availability affects kidney health from development through to disease in a potentially poly(ADP-ribose) polymerase 1 (PARP1)-dependent fashion. Fetuses are frequently exposed to maternal conditions that lower NAD+ abundance including malnutrition and diabetes. These conditions are also associated with reduced fetal nephron endowment (oligonephronia) and adulthood CKD. Little is known about the direct impact of maternal NAD+ on fetal nephrogenesis. My preliminary data in mice show that maternal NAD+ deficiency can cause oligonephronia that can be rescued by maternal supplementation of NAD+ precursors. Published data show that NAD+ dependent PARP1 plays a role in progenitor cell differentiation in multiple cells and tissues during development. Hypothesis: I hypothesize that reduced maternal NAD+ precursor availability may impair fetal nephrogenesis by reducing nephron progenitor cell (NPC) PARP1 activity. Aims: In this proposal, Aim 1 will study how exogenous NAD+ affects NPC differentiation. Aim 2 will assess how cellular compartmentalization of NAD+ influences nephron progenitor cell differentiation using innovative biosensors to characterize how NAD+ biosynthesis compartmental fluxes change through renal development. Aim 3 will elucidate if and how PARP1 mediates the effect of NAD+ on nephrogenesis. The findings from these Aims may provide novel insight into the nutritional aspects of the maternal-fetal interactions that influence renal development and may suggest therapeutic targets for developmental nephron augmentation. Impact: This five-year proposal will provide the training and mentorship necessary for me to develop into an independent investigator studying the metabolism-driven phenomena of renal development, with a specific focus on NAD+. To accomplish this, I have developed a detailed career development plan integrating the outstanding mentorship and scientific environment at UT Southwestern with focused training in developmental biology to successfully transition to independence as an expert at the intersection of metabolism, nutrition, maternal-fetal health, and kidney development.

NIH Research Projects · FY 2026 · 2025-05

Abstract Receptor tyrosine kinases (RTKs) are a large family of single-pass transmembrane cell surface receptors that play key roles in normal cellular processes and are linked with many human diseases. In general, the ligand binding to the extracellular region of RTKs induces receptor dimerizaƟon and acƟvaƟon, but many RTKs cannot be acƟvated simply by ligand-induced dimerizaƟon. In the past 5 years, my lab has performed extensive structural (cryo-EM) and funcƟonal invesƟgaƟons of several special RTKs, including the IR, IGF1R, IRR, c-MET, RET, and MuSK. Each of the RTK stories from my lab individually defines how an important RTK is acƟvated through a unique mechanism. However, many quesƟons regarding the acƟvaƟon and regulaƟon of RTKs remain unanswered. Firstly, to date, none of the cryo-EM structures were able to resolve a complete structure of any RTKs, due to the flexible linkages of the TM with both the ECD and ICD. Therefore, how the conformaƟonal changes are coupled between the extracellular, transmembrane and intracellular regions remain unclear. Secondly, some RTKs are capable of triggering disƟnct downstream pathways when acƟvated by different ligands. These issues of signaling specificity and biased agonism are criƟcal to RTK funcƟon in vivo, but are not understood in physical terms. Thirdly, many RTKs such as TAM, epidermal growth factor receptor (EGFR) and Eph receptors form large clusters upon acƟvaƟon, but the underlying mechanisms are poorly understood. Our future research will be centered on answering these quesƟons, in order to define the general working principles of RTKs. Our innovaƟve approach combines biochemistry, biophysics, cell biology, ligand engineering, and cryo-EM. Finally, understanding the structural basis for the RTK acƟvaƟon and regulaƟon will enable us to design new therapeuƟcs to treat diseases that are caused by malfuncƟons of RTKs.

NIH Research Projects · FY 2026 · 2025-04

PROJECT SUMMARY A significant portion of our behavioral repertoire is initially learned by imitating the behaviors of our parents and other people we interact with while we are young. This is particularly evident when we consider the development of social skills and communicative behaviors like speech and language. Understanding how the brain encodes and retains the memories of social/vocal models that are used to guide accurate imitation is a major scientific challenge. The goal of this research is to identify the cell types and synaptic pathways that encode memories used for learning vocalizations and to map synaptic plasticity mechanisms underlying how these memories are formed in the brain. Studying these memories in the laboratory is complicated by the social nature of their acquisition and the difficulties in isolating discrete and quantifiable imitated behaviors in conventional model species. For example, rodents and non-human primates do not appear to imitate skilled communicative behaviors like vocalizations, but rather communicate using largely innate vocal repertoires. Zebra finches, like people, require social experience from vocal models to form memories used to subsequently guide learning of their courtship song. Unlike some other songbirds, zebra finches do not learn well from passive playback of songs or triggered playback of tutor songs. The study of zebra finch song learning has therefore provided the principal laboratory model for understanding how neural circuits encode memories from social-vocal models that are used to guide precise motor imitation. Research over the last several years demonstrates that the premotor song region HVC is essential for encoding song memories. Yet, neither the neuronal subtypes within HVC nor the presynaptic input pathways necessary for memory encoding have been identified. Using methods for cell- type specific memory manipulation, synaptic connectivity and plasticity mapping, and closed-loop optical inhibition we will define the synaptic pathways encoding song memories and how these memories are synaptically used to guide vocal imitation. In the first aim, we will use optogenetic tools for memory erasure to identify cell subtypes encoding tutor song memories. In the second aim, we will describe the synaptic connectivity and plasticity mechanisms underlying memory formation in these same circuits. In the third aim we will use closed loop silencing of these synaptic pathways to examine the necessity of individual nodes of the circuit in forming vocal memories and vocal imitation. Building on a suite of cutting-edge methods we have developed for use in zebra finches and extensive new data mapping the long-range synaptic connectivity in the songbird brain, this research will identify core circuit mechanisms for forming memories necessary for vocal imitation.

NIH Research Projects · FY 2026 · 2025-04

STING is an ER-associated membrane innate immune protein vital for cancer defense. Despite promising preclinical results, STING agonists have not shown significant success in early clinical trials due to complex signaling, transient activation as a result lysosomal degradation, and detrimental side-effects on healthy tissues. The first Aim of this project is to understand novel lipid-dependent activation and regulatory mechanisms of STING, guiding the development of improved STING-targeting cancer therapies. Our previous research identified PSC7A, a polymer that form nanoparticles that not only can efficiently deliver STING agonists to cells but also can directly bind and activate STING on their own. The PSC7A-activated STING avoids lysosomal degradation seen with the endogenous ligand cGAMP, thereby exhibiting prolonged STING activation and type-I interferon expression, leading to improved antitumor efficacy. The second Aim of this project is to apply cryo-EM to dissect the molecular mechanism underlying PSC7A- induced STING activation. In the third Aim, we will test PSC7A nanoparticles loaded with cGAMP, STING agonists, and specific lipids in various animal tumor models to evaluate their synergistic antitumor effects. Ultimately, this research will facilitate the design of the next-generation STING agonists that precisely control immune signaling, with maximal antitumor immunity but minimal systematic immune-related toxicity in healthy tissues.

NIH Research Projects · FY 2026 · 2025-04

PROJECT SUMMARY The activity-regulated cytoskeletal-associated protein (Arc, also known as Arg3.1) is an immediate early gene product induced by activity/experience and required for multiple modes of synaptic plasticity, including long-term potentiation (LTP), long-term depression (LTD), and homeostatic scaling. It has been implicated in functional and morphological changes in dendritic spines that underlie learning and memory. The best-characterized function of Arc is enhancement of the endocytic internalization of AMPA receptors (AMPARs), a process associated with LTD. Aside from studies showing that Arc binds directly to three elements of the endocytic machinery: the membrane endophilin, dynamin and AP-2, almost nothing is known about the mechanism or regulation of Arc function in AMPAR internalization. We previously reported that Arc palmitoylation is important for mGluR-mediated synaptic weakening, and more recently found that phosphorylation by PKC blocks this modification, suggesting that PKC phosphorylation may serve to limit LTD and/or to enhance LTP. The two PKC phosphorylated serines lie within the endophilin binding motif, raising the possibility that their phosphorylation inhibits the Arc-endophilin interaction. Arc is phosphorylated by the MAP kinase ERK on a serine located within the dynamin and AP-2 binding motifs, again suggesting an inhibitory role for this modification. We recently observed that Arc is an excellent substrate for TAOK2, a kinase that stabilizes the postsynaptic density scaffolding protein PSD95. TAOK2 phosphorylates Arc within its C-terminal domain, the site of interaction with the AMPAR auxiliary subunit, stargazin. Work from others demonstrated that Arc competes with PSD95 for binding to stargazin, suggesting a model in which dissociation of stargazin from PSD95 and association of stargazin with Arc contributes to the delivery of Arc from the PSD to endocytic zones within dendritic spines. We hypothesize that Arc interactions with endophilin, dynamin, AP-2, and stargazin are inhibited upon phosphorylation of Arc by PKC, ERK, and/or TAOK2. In Aims 1 and 2 of this proposal we will use biochemical and biophysical methods and fluorescence-based nanoimaging approaches to characterize the interactions of Arc with endophilin, dynamin, AP-2, and stargazin in vitro and in living heterologous cells. Effects of phosphorylation by PKC, ERK, and TAOK2 on these interactions will be assessed. Based on information obtained in Aims 1 and 2, a subset of interactions and phosphorylation site mutations will be investigated in hippocampal neurons from Arc knockout mice. Synaptic weakening will be examined electrophysiologically and AMPAR internalization will be monitored using immunohistochemical and surface biotinylation assays. We will also measure changes in Arc phosphorylation at sites phosphorylated by the three kinases in response to LTP- and LTD-inducing protocols. Increases and decreases in Arc expression have been linked to numerous cognitive disorders, including Fragile X syndrome, Alzheimer’s disease, and substance abuse. Therefore, we anticipate that elucidation of novel mechanisms of Arc regulation will have therapeutic significance.

NIH Research Projects · FY 2026 · 2025-04

PROJECT SUMMARY Hypertrophic cardiomyopathy (HCM) is the most common genetic cardiovascular disorder. Over 35% of the ~400 mutations identified in HCM patients occur in the motor protein β-myosin. In vitro studies of the R403Q mutant β-myosin revealed hypercontractility of myosin cross-bridge dynamics, including increased ATP turnover and more myosin molecules in the disorganized relaxed sate (DRX) that enables excessive β-myosin contributions to ventricular contractions. When treated with the only FDA-approved HCM-drug mavacamten, the biochemical and structural properties of the isolated mutant β-myosin resembled the WT control. However, direct observation of β-myosin’s conformational alterations due to HCM inducing mutations or mavacamten treatment have yet to be observed in situ. Cryo-electron tomography (cryo-ET) is a powerful imaging technique that enables the visualization of several native cellular macromolecular structures inside intact cells, such as intact myofibrils from cardiomyocytes. However, many of the cryo-ET studies of muscle have been limited to isolated subcellular structures in vitro rather than intact, unperturbed (cardio)myocytes. Additionally, studies often immersed the isolated contractile machinery in conformation inducing buffers before plunge freezing and cryo-ET, possibly further altering the native ratio of myosin active and inactive states. There is a direct need to identify if the isolation procedures utilized to obtain myofibrils result in similar β-myosin structural characteristics found in situ. Further, accurate depiction of native wild type cardiac β-myosin structural features are required to accurately understand how pathological mutations and treatments influence and affect cardiac function and health. To view these contractile proteins within their cellular context, technically difficult high pressure freezing (HPF) and cryo- focused ion beam (cryo-FIB) milling is needed to ensure pristine vitrification of the cells and their organelles. Few labs worldwide have successfully implemented an HFP/cryo-FIB milling/cryo-ET workflow for intact eukaryotic cells and tissues in their native state, including the lab of Dr. Daniela Nicastro, the sponsor of this application. Therefore, I aim to determine the native in situ structure of myofibrils within intact inducible pluripotent stem cell derived human cardiomyocytes using high pressure freezing, followed by cryo-FIB milling to generate thin lamellae that can be imaged by cryo-ET. I will apply this cryo-ET workflow also to characterize structural alterations that occur within sarcomeres due to β-myosin mutations with and without mavacamten treatment. Determining the structural alterations that occur due to mutant β-myosin and mavacamten treatment will provide important insight for future improvement and development of therapeutics able to improve the health of myopathy patients.