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U.S. National Science Foundation

This solicitation addresses issues of translation and diffusion that arise in moving knowledge gained from fundamental learning and education research toward application in PreK-12 STEM classroom practice or leveraging knowledge derived from effective practice toward driving fundamental research. The first goal of this funding opportunity is to encourage the scientific study of theories, frameworks, and models for the translation and diffusion of knowledge, especially between fields and across contexts and levels-of-analysis (e.g., biological to cognitive/socioemotional to behavioral; individual to classroom to broader demographic variables; lab to classroom to school to district). The second goal is to advance or move specific practice, research or scientific discovery in STEM education reciprocally along the research-practice continuum. The Translation and Diffusion (TD) solicitation invites four types of proposals: Research on Translation or Diffusion proposals request funding to conduct scholarship that will advance the sciences of translation or diffusion of fundamental research knowledge toward PreK-12 formal STEM education practice by developing or refining theories, frameworks, or models (or adapting those from other domains) and conducting related research. Such proposals may also address the leveraging of effective classroom practices toward the enrichment of foundational research, constructs and models. We note that bi-directional movement across boundaries is a mutually beneficial reciprocal process. Proof-of-Concept Research proposals request funding to explore the feasibility and viability of particular knowledge or products generated from STEM education research toward advancing practice in formal PreK-12 settings (even if it is still basic or applied research and development rather than implementation). The goals are to facilitate the process by which the promise that the initial insight holds for research and practice can be realized. The outcome of such a project would lay the methodological, theoretical, empirical, design, or social foundation for conducting systematic work at the next stage of development or at the next level of analysis. Empirical and theory-building efforts to adapt initial insights from research or practice across significantly different contexts, populations, domains, and levels-of-analysis are also welcome. Synthesis proposals critically integrate the current state of knowledge on a particular topic relevant to translation and diffusion in formal PreK-12 STEM education. Such proposals should include the state of the knowledge across disciplinary communities and across relevant literatures, identify the lacunae in STEM education knowledge, and, where appropriate, lay out thenext steps for future research and development. Conference / Workshop proposals relevant to the call are also welcome.

NHLBI - National Heart Lung and Blood Institute

Project Summary/Abstract Despite surgical advancement and endovascular device innovations, endothelial injury remains a fundamental barrier in vascular surgeries, contributing to post-operative complications such as vein graft failure, restenosis, and thrombosis. Current surgical approaches offer no regenerative solution to actively protect endothelial health during or after interventions. To address this gap, we have developed a bioengineered mitochondrial therapy platform that leverages lipid-polymer-peptide surface functionalization to enable targeted mitochondria delivery, enhanced cellular uptake, and integration into the host mitochondrial network. Mitochondrial dysfunction is a key driver of impaired endothelial repair following vascular surgeries, yet mitochondrial transplantation has not been adopted clinically due to challenges in storage stability, scalability, and compatibility with surgical workflows. Our preliminary studies demonstrate successful surface engineering of stem cell–derived mitochondria with robust uptake and functional rescue in dysfunctional endothelial cells. In vivo, we show that engineered mitochondria were selectively uptake by target vessels during both intra-operative (ex vivo vein graft storage and arterial intraluminal infusion) and post-operative stages (intravenous injection) upon vascular surgeries. To further establish the translational potential of bioengineered mitochondrial therapy, we propose the following studies. Aim 1 will optimize mitochondrial source selection, storage formulation, and surface modification parameters to ensure translational feasibility. Aim 2 will validate the therapeutic efficacy and biodistribution of the optimized system in two highly clinically relevant in vivo models of open (vein graft bypass) and endovascular surgeries (balloon angioplasty). This interdisciplinary project integrates surgical sciences, bioengineering, translational medicine, and device development to establish a next-generation therapeutic approach for vascular surgeries. By advancing a procedural-compatible, device-enabled delivery system, this work directly addresses a critical need in vascular surgery by developing translatable tools and technologies to improve surgical outcomes in millions of patients with cardiovascular diseases.

NICHD - Eunice Kennedy Shriver National Institute of Child Health and Human Development

ABSTRACT Transposable elements (TEs) make up over half of the human genome and are increasingly recognized as key regulatory sequences of gene expression in development and disease. While cellular intrinsic transcriptional and epigenetic mechanisms controlling TE sequences are well-documented, whether and how TEs response to external microenvironmental signals, particularly mechanical forces, remains unexplored. This represents a critical gap in understanding both TE regulation and genome–environment interactions. Our preliminary data reveal that several TE families, including LTR7 from the primate-specific HERV-H family, function as Mechano-Response Enhancer Elements (MREEs) that regulate gene expression and human pluripotent stem cell fate in response to mechanical stimuli. Additional preliminary findings support our central hypothesis that TEs act as MREEs by modulating human genes and cell fate of human pluripotent stem cells through a mechanism at least partially governed by the key mechano-effector YAP, which regulates TEs’ local epigenetic activity and facilitates their long-range chromatin looping with target genes in response to mechanical changes. To test this model, we propose three specific aims: 1) determine how mechanical signals regulate the local chromatin activity of TE MREEs; 2) elucidate how mechanical cues mediate long-range chromatin interactions between TE MREEs and their distal target genes; 3) delineate the mechanism by which TE MREE modulating human embryonic stem cell fate. Our objective is to rigorously establish the novel concept that TEs function as MREEs, uncover their underlying molecular mechanisms, and assess their biological significance in regulating developmental genes and hESC fate. The expected outcome will not only shift the current paradigm of mechanobiology from protein- coding genes to non-coding regulatory elements such as TE but also improve our understanding of TE regulation and genome-environment interactions in health and disease.

NICHD - Eunice Kennedy Shriver National Institute of Child Health and Human Development

Summary: The negative effects trauma exposure, such as those that promote PTSD, can be passed to offspring. While the environment in which affected parents raise their offspring clearly plays a role, a significant genetic component also exists. However, some of this genetic influence, typically revealed by twin studies, may stem from epigenetic inheritance; as implied by studies in male rodents, where trauma-specific changes in sperm miRNA content can lead to trauma-specific behavioral changes in all offspring. Evidence for epigenetic inheritance in humans is mainly epidemiological, but recent studies have revealed stress-induced sperm miRNA changes consistent with this idea. For example, we found that men with high Adverse Childhood Experiences (ACEs), whose children can be negatively affected, show reduced sperm levels of miR-34/449 family members; mirroring findings in mice exposed to chronic social instability (CSI) stress across generations. In mice, these sperm miRNA changes persist in preimplantation embryos post-fertilization, altering early embryonic gene expression that leads to elevated anxiety and impaired sociability in female offspring as well as reduced levels of sperm miR-34/449 in males. Our new data driving this proposal reveals another example. The degree of men’s exposure to adult trauma, as assessed by the Trauma History Questionnaire (THQ), which measures PTSD risk, correlates with the levels of miRNAs 532, 361, 375, and 491 in their sperm. The highest THQ scores are associated with 4- to 130-fold increases in these levels. In contrast, these miRNA changes do not correlate with ACE scores and miR-34/449 does not correlate with THQ score. Notably, miRNAs-532 and 375 are two of the 9 sperm miRNAs whose enhanced levels in male mice mediate how chronic variable (CV) stress leads to a suppressed HPA axis response in offspring, a trait linked to mental health disorders. This proposal investigates how elevated levels of sperm miRNAs-532,361,375 and 491 in men with high THQ scores might affect their offspring using the mouse model that previously demonstrated how elevated levels of sperm miRNAs transmit the stress-related effects of paternal CV stress across generations. Aim 1 tests how injecting THQ-associated miRNAs, at levels found in sperm of men exposed to high levels of trauma, into mouse zygotes affects the phenotypes of resulting offspring. Aim 2 will begin to reveal how these injected miRNAs lead to phenotypic changes revealed in Aim 1 by i) identifying gene expression changes induced by them in mouse blastocysts, and ii) looking for similarities to those occurring in discarded blastocysts from IVF procedures using men’s sperm with elevated THQ associated miRNAs derived from an ongoing, independently funded project. This study has the potential to: a) provide strong support for using mouse models to understand epigenetic inheritance in humans; and b) drive future research showing that a significant portion of inherited susceptibility to mental health disorders arises from epigenetic inheritance, and how to reverse it before fatherhood.

U.S. National Science Foundation

The Tribal Colleges and Universities Program (TCUP) provides awards to federally recognized1 Tribal Colleges and Universities, Alaska Native-serving institutions, and Native Hawaiian-serving institutions to promote high quality science (including sociology, psychology, anthropology, linguistics, economics and bioeconomics, statistics, and other social and behavioral sciences; natural sciences; computer science, including, but not limited to, artificial intelligence, quantum information science, and cybersecurity), technology, engineering and mathematics (STEM), STEM education, research, and outreach. Support is available to TCUP-eligible institutions (see the Additional Eligibility subsection of Section IV of this solicitation) for transformative capacity-building or community engagement projects through Instructional Capacity Excellence in TCUP Institutions (ICE-TI), Targeted STEM Infusion Projects (TSIP),TCUP for Secondary and Elementary Teachers in STEM (TSETS), TCU Enterprise Advancement Centers (TEA Centers), Cyberinfrastructure Health, Assistance, and Improvements (CHAI), and Preparing for TCUP Implementation (Pre-TI). Collaborations led by TCUP institutions that involve non-TCUP institutions of higher education are supported through TCUP Partnerships, with the participation of other NSF programs to support the work of non-TCUP institutions. Finally, research studies that further the scholarly activity of individual faculty members are supported through Small Grants for Research (SGR). Through the opportunities highlighted above, as well as collaborations with other National Science Foundation (NSF) divisions and directorates, and other organizations,TCUPaims toincrease Native individuals' participation in STEM careers, improve the quality of STEM programs atTCUP-eligible institutions, and facilitate the development of a strong STEM enterprise in TCUP institutions' service areas. TCUP supports transformative capacity-building, community engagement, or research projects at TCUP-eligible institutions through the following funding tracks: Instructional Capacity Excellence in TCUP Institutions (ICE-TI) projects provide support to design, implement, and assess comprehensive institutional improvements in STEM education and research capacity at TCUP-eligible institutions of higher education. By strengthening STEM education and STEM education research, successful projects will increase the number of STEM students and improve the quality of their preparation. ICE-TI projects create and/or adapt and assess innovative models and materials for teaching and learning in STEM, embody knowledge about how students learn most effectively in STEM teaching and learning activities, and bring STEM disciplinary advances into the undergraduate or graduate experience. The objective of this strand is to expand STEM degrees offered by TCUP-eligible institutions or significantly enhance instructional approaches. Targeted STEM Infusion Projects (TSIP) support the attainment of a short-term, well-defined goal to improve the quality of STEM education at an eligible institution. Targeted STEM Infusion Projects could, for example, enhance academic infrastructure by systematically adding traditional knowledge to the scope or content of a STEM course, updating curricula, modernizing laboratory research equipment, developing and delivering professional development for K-12 STEM educators, or improving the computational infrastructure.The objective of this strand is to expand STEM degrees or significantly enhance instructional approaches. TCUP for Secondary & Elementary Teachers in STEM (TSETS) supports in-service professional development in STEM disciplinary or STEM education content and/or research for K-12 STEM teachers in the relevant service area. Examples of project activities include, but are not limited to, professional development involving seminar series and engagement in STEM instruction and content during the academic year, structured series of summer intensive workshops and trainings, and summer research opportunities. The objective of this strand is to broaden the instructional capacity for STEM in the K-12 workforce and thereby to the entire community, and to build the capacity for STEM disciplinary or education research among participating educational professionals. TCU Enterprise Advancement Centers (TEA Centers) coalesce the STEM and/or STEM education expertise into a team, designed to support and promote the STEM goals, needs, aspirations, or interests of the chartering reservation or tribe(s). TEA Centers may address a critical tribal or community need or focus on a realm of research or design that is beyond the scope of individual research grants or that is of interest to multiple tribes. The objective of this strand is to build on the capacity developed through prior TCUP support and apply expertise to collaborations with communities in the institution s service area, or nationally. The Cyberinfrastructure Health, Assistance, and Improvements (CHAI) strand supports projects at TCUP-eligible institutions of higher education to upgrade the cyberinfrastructure necessary to conduct, expand, manage and administer STEM programs of study, including research. The objective of this strand is to equip TCUP institutions to meet the demands of virtual instruction, advanced computing, and data science opportunities. Preparing for TCUP Implementation (Pre-TI) provides support for activities that prepare an institution for Implementation-level projects.Consequently, they are available only to TCUP-eligible institutions of higher education that have never received TCUP support, have not received TCUP support within the previous five years, or are embarking on a significantly novel STEM strategic plan. Examples of supported activities include completing an institutional assessment of its current STEM instructional capacity, or engaging in conversations necessary to formulate a shared vision of what that capacity should be and how to achieve it. Pre-TI awards can support staff and faculty release time, travel, stakeholder gatherings, and associated administrative costs.The objective of this strand is to conduct self-studies and formulate strategic plans for the development of STEM instructional programs of study. The TCUP Partnerships strand provides support for collaborations that will improve TCUP institutions' instructional and research capacity in STEM fields supported by NSF; attract, retain, and support TCUP students in internships and research endeavors deemed to be necessary for a complete curriculum offering; and engage partner universities to provide an academic grounding and a successful transition for students who wish to study or attain degrees in STEM fields supported by NSF. TCUP Partnerships broaden the number of scientific disciplines available to students at TCUP institution through collaborations with non-TCUP institutions. Active Pre-Engineering Education Collaboratives or Partnerships in Geoscience Education awards are not affected by this revision. The objective of this strand is the development, through instructional and research capacity-building, of academic and career pathways for TCUP students through supporting collaborative projects between and among TCUP and non-TCUP institutions. Interested teams of collaborators for which a TCUP institution serves as lead should contact the TCUP program directors. Support for non-TCUP partners must be obtained from other NSF programs, which follows the procedures of the prior Partnership strands. Small Grants for Research (SGR) strand support STEM or STEM Education faculty members at TCUP-eligible institutions to initiate or pursue research projects or programs that may include undergraduate or graduate student engagement. Awards are intended to help further the faculty member's research capability and effectiveness; improve research and teaching at his or her home institution and create and study new models and innovations in STEM teaching and learning. International research or collaborations are strongly encouraged. TCUP students may seek support for international research opportunities under the guidance of a TCUP STEM or STEM education faculty member and an international research collaborator. These awards are particularly appropriate as a means of recruiting and retaining highly qualified scientists, engineers, and educators at TCUP-eligible institutions. The objective of this strand is to support faculty research and professional development that build research capacity at TCUP institutions. [1] Executive Order 13021 defines Tribal Colleges and Universities ("tribal colleges") as those institutions cited in section 532 of the Equity in Educational Land-Grant Status Act of 1994 (7 U.S.C. 301 note), and other institutions that qualify for funding under the Tribally Controlled Community College Assistance Act of 1978, (25 U.S.C. 1801 et seq.), as well as Navajo Community College as authorized in the Navajo Community College Assistance Act of 1978, Public Law 95-471, Title II (25 U.S.C. 640a note). The term "Alaska Native-serving institution" means an institution of higher education that is an eligible institution under section 1058(b) of the Higher Education Act; and that, at the time of submission, has an undergraduate enrollment that is at least 20 percent Alaska Native students. The term "Native Hawaiian-serving institution" means an institution of higher education that is an eligible institution under section 1058(b) of the Higher Education Act; and that, at the time of submission, has an undergraduate enrollment that is at least 10 percent Native Hawaiian students. Most TCUP-eligible institutions of higher education are two-year or community colleges. See the Who May Submit Proposals section in this solicitation for further details.

NICHD - Eunice Kennedy Shriver National Institute of Child Health and Human Development

PROJECT SUMMARY Placental aging is the programmed process of progressive functional decline and tissue maturation that occurs throughout the course of pregnancy, particularly in the later stages. While this is a normal feature of development, accelerated or pathological placental aging triggered by stress can impair placental function and lead to serious complications, including stillbirth, preeclampsia, fetal growth restriction, and preterm birth. Understanding mechanisms driving premature placental aging is critical for developing strategies to improve pregnancy outcomes. This proposal tests the hypothesis that precocious trophoblast differentiation leads to exhaustion of the trophoblast stem cell (TSC) pool, triggering premature placental aging and disease. TSCs maintain placental homeostasis by regenerating the syncytiotrophoblast – the multinucleated epithelial layer that mediates nutrient and gas exchange at the maternal-fetal interface. Loss of TSC self-renewal or differentiation under stress may compromise placental regenerative capacity and contribute to aging and failure. Our preliminary data identify the transcription factor CEBPB as a key regulator of the stress response and TSC differentiation. Conversely, trophoblast-associated microRNAs, including the murine miR-290~295 cluster and its human ortholog miR- 371~373, help preserve TSC identity by supporting stem cell self-renewal and metabolism. Loss of miR-290 in mice leads to premature depletion of the TSC pool, the accumulation of aging markers in the placenta, and stillbirth – linking early stem cell dysfunction with placental failure. The overall goal of this project is to define the molecular and metabolic pathways that govern TSC differentiation and placental aging. Aim 1 will characterize stress-induced TSC differentiation and senescence, focusing on the regulatory roles of CEBPB and miR- 371~373. Aim 2 will determine how TSC depletion contributes to placental aging and stillbirth in miR-290 knockout mice, using transcriptomic and histologic approaches. Aim 3 will investigate the role of metabolic reprogramming in TSC fate and aging, and test whether metabolic interventions can preserve stemness under stress. By linking TSC biology with placental aging and adverse outcomes, this work provides a conceptual framework that opens avenues for therapeutic innovation. Applying principles from aging biology to the placenta may enable repurposing of existing anti-aging strategies to reduce placental dysfunction and prevent the most devastating consequence—stillbirth. This project directly addresses the goals of NOSI NOT-HD-23-021: The Road to Prevention of Stillbirth.

NIGMS - National Institute of General Medical Sciences

Primary cilia play a pivotal role in human health, acting as signaling hubs to sense extracellular cues such as odorants, metabolites, light, neurotransmitters, and more. Defects or failure of these signaling hubs to function leads to developmental disorders, immune dysfunction, diabetes, obesity, Parkinson’s disease, cancer, and other pathologies. An early focus of this grant is the generation of better tools to dissect primary cilia function and mechanism, such as a protocol for synchronized ciliogenesis in human retinal pigmented epithelial (RPE) cells coupled with microscopy and shotgun/phosphoproteomic mass spectrometry, to dissect signaling events temporally and spatially. While mapping ciliogenesis via immunofluorescence with known markers of the stages, one protein that emerged as a strong candidate for regulating interpathway communication was TULP3, a 50kDa ciliary protein whose primary known function is in driving the import of ciliary GPCRs through unclear mechanisms. A combination of synchronization and classical cell biological approaches were used to uncover novel phenotypes revealing temporal and spatial timing of TULP3’s function in GPCR traffic as well as new functions in ciliogenesis, downstream of TTBK2 recruitment but before axoneme protrusion. Patient mutations were identified from the use of GWAS databases to probe for links between protein function and human health. Constitutive expression of these TULP3 mutants in TULP3 KO background generated new tools to perturb select functions for TULP3, which is especially powerful for probing function-specific binding domains and partners. This proposal tests the following hypotheses: TULP3 (i) regulates receptor traffic prior to cargo arrival at the basal body and coupling to IFT machinery to mediate receptor entry into cilia and (ii) licenses axonemogenesis. Furthermore, primary cilia perform two different functions in lymphoid tissues: regulating fate change of hematopoietic stem cells into lymphocytes in the bone marrow, and in the lymph node facilitating lymphocyte maturation. These hypotheses will be tested in the following aims: Aim 1- Determine the mechanism by which TULP3 mediates membrane receptor transport into cilia. Aim 2- Identify the function and mechanism of TULP3 in ciliogenesis. Aim 3- Uncover the function of primary cilia and TULP3 in the generation of white blood cells. These aims will be addressed using cell culture and mouse model systems to probe TULP3 function and its role in immunity in both in vitro and in vivo contexts. The success of any and all of these aims will provide novel insight into key mechanisms driving ciliary function and reveal cellular contexts for how disrupted immune function arises. The study of primary cilia in the lymphatic system is largely uncharted territory. As a result, these studies are expected to open new fields of investigation into mechanisms of immune regulation.

NCI - National Cancer Institute

PROJECT SUMMARY Metastasis is the leading cause of mortality among patients diagnosed with solid tumors. In this regard, identifying common mechanisms within primary, premetastatic and metastatic niches that contribute to progression of solid tumors represents a significant need. The Lipocalin-2 gene (LCN2, neutrophil gelatinase- associated lipocalin or NGAL) encodes a cytosolic and secreted protein (Lcn2) that regulates receptor trafficking, innate immune responses, inflammation, microbiome dynamics and iron homeostasis. Previous work from our group and others has shown that Lcn2 can promote stemness and tumorigenesis in models of solid tumor progression. However, uncertainty persists about the stage or stages at which LCN2 exerts tumorigenic effects and whether the mechanisms of LCN2 action vary based upon stage or tumor location. In this regard, we have recently performed unbiased single-cell spatial proteomic and transcriptomic screens of breast and pancreatic cancer patient samples leading us to define tumor cell autonomous and non-atonomous roles for Lcn2/LCN2 in promoting solid tumor progression. Based upon these findings, the central hypothesis of this proposal is that Lcn2 promotes FGFR2 signaling and adaptive anti-inflammatory immunity to support solid tumor progression. The overall objective of this proposal is to determine the spatiotemporal dynamics of Lcn2- governed tumor cell autonomous and non-autonomous mechanisms driving metastatic progression of solid tumors. Our rationale for pursing this work is that understanding these mechanisms will position Lcn2 as a biomarker for immune therapy resistance and target for improving immune therapy success in immunologically cold tumors. To test the central hypothesis, we propose to specific aims that will (I) identify the tumor cell autonomous signaling mechanisms governing Lcn2-dependent FGF2-induced tumor cell invasion and (II) define the intercellular communication mechanisms by which Lcn2 reduces inflammation and enriches regulatory T cells within the metastatic niche. The basis of this project is conceptually innovative and employs technically innovative transgenic, pharmacology, cell cycle reporter, protein reporter tagging, single-cell spatial omics, spinning disc time-lapse confocal and CRISPR methods in combination with preclinical allo- graft/xenograft models of solid tumors and hetero-multicellular cancer spheroid cultures. This work is expected to (1) provide exceptional research opportunities for undergraduates while enhancing the research environ- ment at Baylor University and (2) elucidate targetable mechanisms that govern solid tumor progres- sion/metastasis. As such, the proposed studies will have a positive impact on Baylor student success and identify novel treatment strategies to improve cancer outcomes.

NCI - National Cancer Institute

PROJECT SUMMARY Acute myeloid leukemia (AML) is an aggressive, devastating cancer with limited treatment options. AML progresses rapidly and presents significant treatment challenges due to its immunosuppressive tumor microenvironment , which impairs immune cell function. Group 1 innate lymphoid cells (ILCs), including natural killer (NK) cells and ILC1s, play key roles in immunity. ILC1s reside in tissues and were initially believed to function primarily by secreting cytokines such as IFN-γ, TNF-α, and GM-CSF. However, their anti-tumor activity has been largely unknown. In 2022, we addressed this gap and published our discovery in Nature Immunology as a cover story. We found that ILC1s isolated from AML patients are functionally impaired, whereas ILC1s from healthy mice are significantly more potent. Healthy ILC1s induce the death of leukemia stem cells (LSCs), block LSC differentiation into leukemia progenitor cells, and promote the transition of LSCs into non-leukemic lymphoid progenitors. Mechanistically, normal ILC1s target LSCs by secreting IFN-γ and engaging receptor-ligand interactions (e.g., DNAM-1–CD155 and IL-7 receptor–IL-7). Despite identifying key features of ILC1s and their role in inhibiting LSCs, important questions remain unanswered. It is still unclear how ILC1s develop in vivo under normal or AML conditions, and the mechanisms through which ILC1s induce LSC death and differentiation in humans are largely unexplored. Moreover, the therapeutic potential of ILC1s remains unknown. We hypothesize that ILC1s possess strong anti-LSC activity and unique developmental pathways, offering a novel approach to control or treat AML and potentially prevent its relapse. The goals of this project are to elucidate the mechanisms of ILC1 anti-tumor activity, characterize their developmental pathways, and explore their therapeutic applications. In Aim 1, we will dissect the mechanisms by which ILC1s induce LSC death (e.g., via pyroptosis) and drive M1 polarization of LSC-differentiated myeloid cells in humans. In Aim 2, we will characterize ILC1 developmental pathways in both normal and AML conditions. Leveraging our expertise in developing adoptive cellular therapies, including chimeric antigen receptor (CAR) NK cells for AML, in Aim 3, we will study novel FLT3-targeting CAR ILC1s that we generated. FLT3 is highly and selectively expressed on AML blasts and LSCs, making it an ideal target. We will generate allogeneic, off-the-shelf, ready-to-use FLT3- CAR ILC1s from umbilical cord blood CD34⁺ cells or by converting NK cells into ILC1s, which we demonstrated. These CAR ILC1s will be tested for their anti-AML efficacy in preclinical models and compared to unmodified ILC1s. Additionally, we will combine ILC1s or FLT3-CAR ILC1s with NK cells and an FDA-approved tyrosine kinase inhibitor, which upregulates FLT3 expression on AML cells. Finally, we propose to reprogram endogenous ILC1s by treating them with IL-7 to enhance their activity. A deeper understanding of ILC1 development and function, anticipated through the completion of this study, holds significant promise. The knowledge gained could lay the groundwork for diverse therapeutic strategies that have the potential to reduce mortality in AML patients.

U.S. National Science Foundation

The NSF Research Traineeship (NRT) program seeks proposals that explore ways for graduate students in research-based master s and doctoral degree programs to develop the skills, knowledge, and competencies needed to pursue a range of STEM careers. The program is dedicated to effective training of STEM graduate students in high priority interdisciplinary or convergent research areas, through a comprehensive traineeship model that is innovative, evidence-based, and aligned with changing workforce and research needs. Proposals are requested that address any interdisciplinary or convergent research theme of national priority, as described in section II.D below. The NRT program addresses workforce development, emphasizing broad participation, and institutional capacity building needs in graduate education. The program encourages proposals that involve strategic collaborations with the private sector, non-governmental organizations (NGOs), government agencies, national laboratories, field stations, teaching and learning centers, informal science centers, and academic partners. NRT especially welcomes proposals that reflect collaborations between NRT proposals and existing NSF Eddie Bernice Johnson Inclusion across the Nation of Communities of Learners of Underrepresented Discoverers in Engineering and Science (INCLUDES) Initiative, Research Experiences for Undergraduates (REU), Louis Stokes Alliances for Minority Participation (LSAMP), NSF Scholarships in Science, Technology, Engineering, and Mathematics (S-STEM), and NSF STEM Ed Organizational Postdoctoral Fellowship program (STEM Ed OPRF) projects, provided the collaboration will strengthen both projects. Researchers at minority serving institutions and emerging research institutions are strongly encouraged to submit proposals. Collaborations between NRT proposals and existing NSF INCLUDES projects should strengthen both NRT and INCLUDES projects.

NIAMS - National Institute of Arthritis and Musculoskeletal and Skin Diseases

ABSTRACT Epidermis of the skin undergoes continuous self-renewal through a tightly regulated balance of keratinocyte proliferation and terminal differentiation. Disruption of this balance is characteristic of inflammatory skin disorders such a psoriasis, atopic dermatitis, and neutrophilic dermatoses. The etiologies of these skin disorders are complex and heterogeneous, as are the needs for treatments. Our long-term goal is to elucidate how dysregulation of K63-Ub-mediated signal transduction pathways in keratinocytes contribute to skin inflammation. Towards this end, our recent studies have focused on UBE2N, a ubiquitin conjugase that forms heterodimers with an essential noncatalytic partner, UBE2V1 or UBE2V2, to specifically catalyze K63-Ub of target proteins. We demonstrate that conditional knockout of Ube2n in mouse keratinocytes induces psoriasis- like inflammatory skin lesions with a raised and scaly appearance. Transcriptomic and histological analyses identified a diminished epidermal stem cell compartment, a thickened epidermal spinous layer, and an increased infiltration of myeloid-skewed immune cells. This is correlated with increased expressions of myeloid cell chemokines such as CXCL1 and CXCL2 and IL1 family cytokines in keratinocytes and infiltrating myeloid cells. Oral delivery of the small molecule inhibitor of IRAK1/4, common mediators of the IL1R and TLR signaling pathways, alleviated immune infiltration and epidermal defects of the mutant skin. These data highlight a key role for UBE2N in regulation of epidermal and cutaneous immune homeostasis. In line with these animal data, recent GWS studies demonstrate a causal association between UBE2V1 polymorphism and psoriasis. Together, these data support the hypothesis that UBE2N partners with UBE2V1 to restrain keratinocyte recruitment of myeloid cells through suppression of IL1 and CXCL1/2-mediated inflammatory crosstalk between keratinocytes and myeloid cells. We propose 3 specific aims to: 1) validate the importance of UBE2N catalytic function and the role of UBE2V1 in epidermal homeostasis and cutaneous immune homeostasis, 2) determine the contribution of the IL1 signaling pathway in UBE2N-null skin inflammation, and 3) assess the utility of CXCL1/2 receptor antagonists in mitigating neutrophilic dermatosis. We will utilize conditional genetic animal models along with the cutting-edge techniques of single cell transcriptomics and global proteomics to comprehensively analyze mechanistic aspects of UBE2N/UBE2V1-mediated K63-Ub in cutaneous inflammation and therapeutic targeting. Results of these studies will reveal novel mechanisms of epidermal and cutaneous immune homeostasis, as well as insights for therapeutic development.

NIDA - National Institute on Drug Abuse

PROJECT SUMMARY Cocaine use disorder (CUD) is highly comorbid in people with HIV (PWH) and can accelerate infection, alter neuropathology, and exacerbate cognitive decline despite antiretroviral therapy (ART). Many of these effects are due to the infection and dysregulation of CNS-associated myeloid cells, especially microglia, which comprise a significant reservoir in this compartment. However, the precise mechanisms by which cocaine (Coc) dysregulates microglia to enhance HIV infection are unclear, partly due to the lack of translationally relevant human microglial models suitable for mechanistic evaluation of Coc-mediated changes in viral dynamics. Classically, Coc has been thought to act by blocking dopamine transporter (DAT) activity, exposing microglia to aberrantly high dopamine concentrations. Our data show that dopamine can increase HIV infection and inflammation in microglia and other myeloid cells. However, recent data show that Coc has other mechanisms of action beyond the modulation of dopaminergic tone, involving the ER protein sigma1 (σ1), which has diverse cellular functions including the modulation of cellular stress pathways such as the unfolded protein response (UPR). Viruses, including HIV, can exploit the UPR to amplify stress-induced protein production in the host cell, enhancing viral replication. Our preliminary studies indicate that Coc’s effects on σ1 may drive a Coc-mediated increase in HIV infection in microglia, potentially through increased stress response and independent of dopamine’s effects. My preliminary data show that both Coc and σ1 agonists increase HIV replication in human inducible pluripotent stem cell (iPSC)-derived microglia (iMg). These effects are blocked by σ1 antagonism but not by inhibition of DAT or dopamine receptors. We also show increased σ1 protein expression and recruitment to the ER/nuclear envelope space in HIV-infected iMg treated with Coc, and preliminary single-cell RNAseq data suggest changes in the UPR. Therefore, we hypothesize that Coc-mediated activation of σ1 increases HIV infection of microglia via activation of the UPR. In Aim 1, we will test the involvement of σ1 in driving Coc-mediated changes in HIV infection of iMg using pharmacological and genetic modulation, and we will also confirm the absence of dopaminergic involvement. We will assess changes in viral dynamics using AlphaLISA and immunofluorescence (IF) high-content imaging. In Aim 2, we will test the hypothesis that Coc induces greater σ1 activity in the presence of HIV infection utilizing confocal and high-content IF imaging of σ1 subcellular localization in cellular compartments like the nuclear envelope, ER, and mitochondria-associated ER membrane. Movement of σ1 to these compartments is a feature of σ1 activation. In Aim 3, we will use single-cell RNAseq to test the hypothesis that Coc-induced σ1 activity drives increased HIV infection in iMg via upregulation of UPR genes. The results from these experiments will not only define novel interactions between HIV and σ1 that could reveal new antiretroviral targets but will also broadly inform on the role of σ1 in microglia and potentially identify biomarkers for prevention strategies against CUD and its associated comorbid diseases.

NHLBI - National Heart Lung and Blood Institute

Abstract: The adult zebrafish is capable of rapid regenerative processes in response to cardiac damage that result in a fully functional and scarless heart. This robust regenerative response is controlled by pre-existing populations of cardiomyocytes that de-differentiate prior to proliferation in response to injury; however, the full extent of intercellular signaling that facilitates the injury-response of these cardiomyocytes remains unknown. A subpopulation of cardiomyocytes in the zebrafish heart are derived from the cardiac neural crest (CNC), a highly migratory stem cell population in the early vertebrate embryo. These neural crest-derived cardiomyocytes (NCCMs) may contribute extensively to regeneration in the adult heart by reactivating developmental gene networks, including those controlling morphogenesis in the early embryo. In particular, the cxcr4b/cxcl12a chemokine signaling pathway is known to be essential for CNC migration in the developing embryo and regulates aspects of migration in the regenerating heart. To assess the roles of cxcr4b/cxcl12a in relation to the CNC during cardiac regeneration in the adult, I will conduct mRNA expression and localization analyses via hybridization chain reaction (HCR), in vitro migration assays of primary cardiomyocyte cultures, and create transgenic fish lines for the conditional overexpression and knockout of cxcl12a and cxcr4b, respectively, during cardiac regeneration in the adult. I will also analyze existing cardiac ATAC-seq data to identify and validate putative damage-responsive enhancer regions that drive chemokine gene expression. Identifying how migratory mechanisms guide cardiomyocytes will assist in discovering therapeutic approaches aimed at stimulating human hearts with a regenerative capacity.

NHLBI - National Heart Lung and Blood Institute

PROJECT SUMMARY/ABSTRACT Mutations in telomerase cause bone marrow failure in patients suffering with dyskeratosis congenita and other associated telomere biology disorders. While mutations in these patients are found in different components of telomerase, mutations in genes that regulate the processing of the RNA component of telomerase, TERC, are the most prevalent. Due to a lack of adequate models and intrinsic difficulties in studying human telomerase in physiologically relevant cells, the molecular pathways that control TERC biogenesis and decay during hematopoiesis remain largely unknown. Progress in the field has been hampered by species and even cell-type specific differences in telomerase biology that limit our understanding of the molecular mechanisms leading to the disproportionate role of TERC in hematopoietic failure when compared to other components of telomerase. A better understanding of the molecular regulation of TERC biogenesis and function in hematopoietic cells is essential for development of novel alternatives for patients, which remain without a cure. The focus of this proposal is to use different in vitro and in vivo approaches to decipher molecular pathways controlling TERC biogenesis and decay in blood cells, as well as the function of TERC during erythroid, myeloid and lymphoid development. We have developed unique models, including targeted hematopoietic differentiation of human pluripotent stem cells, transplantation of primary CD34+ human stem cells into sub-lethally irradiated mice, and studies in primary patient samples, that will allow a complete analysis of the pathways regulating TERC decay and function during hematopoietic development. For that, two specific aims are proposed that will both identify novel regulators of TERC decay in blood cells, as well as specific functions of TERC in the hematopoietic system. Aim 1 will determine the role of novel, recently identified 3'- end RNA deadenylases to TERC processing in the blood, and to which extent different RNA deadenylases prevent TERC degradation by the exosome. We will complement these experiments with the identification of the molecular effectors of a novel route for TERC decay, triggered by differential TERC capping on its 5'- end, and mediated by trafficking to the cytoplasm. We will investigate if modulation of these different pathways can rescue hematopoietic development in telomerase mutants. Aim 2 will investigate novel functions of TERC outside telomerase that can explain the disproportionate role that mutations that affect TERC levels show in bone marrow failure. We have created unique cellular systems where we can uncouple TERC expression from telomere length, and will utilize them during hematopoietic differentiation to study direct functions of TERC on DNA damage and regulation of hematopoietic gene expression programs. These studies will determine the molecular mechanisms controlling TERC decay and function in hematopoietic cells. Our unique cellular tools, combined with our expertise in telomerase, RNA decay, and stem cell biology puts us in an ideal position to make a significant impact in this field.

NIA - National Institute on Aging

PROJECT SUMMARY This NIH F30 grant proposal investigates how somatic mutations in hematopoietic stem cells (HSCs) contribute to aging-related diseases through clonal expansion of mutated blood cells. This phenomenon, called clonal hematopoiesis, becomes more common as individuals age, affecting over 10% of in people over 60 years old. This study focuses on mosaic chromosomal alterations (mCAs), large-scale mutations that are under-studied compared to smaller mutations like clonal hematopoiesis of indeterminate potential (CHIP). mCAs are linked to lymphoid malignancies and infection susceptibility due to lymphoid-biased differentiation, while CHIP correlates with cardiovascular diseases and myeloid malignancies through myeloid-biased differentiation. I hypothesize that mCA expansion is determined by individual factors rather than mCA genetic change and that clones with greater expansion rates will have increased disease risk. Aim 1 examines the influence of mCA characteristics and environmental factors on clonal expansion rates using longitudinal blood samples from 30,000 individuals in Vanderbilt’s BioVU genomic and clinical biobank. Using longitudinal mCA trajectories, I will quantify the contribution of the mCA mutation and individual characteristics (e.g., age, sex, BMI, smoking, type 2 diabetes, lipoprotein levels) to clonal expansion rate and build a predictive model for mCA clonal expansion. My working hypothesis for Aim 1 is that mCA expansion varies widely among individuals with the same mCA and thus modifiable lifestyle exposures are major contributors to clonal expansion rate. The longitudinal samples in BioVU will not be sufficient to test genetic and phenotypic associations with clonal expansion rate. Therefore, Aim 2 expands the study to detect mCAs in > 1 million individuals across various genomic biobanks with single blood draws (i.e., NHLBI TOPMed, NIH All of Us, UK Biobank, and BioVU). To determine mCA clonal expansion rate from a single timepoint, I will apply Passenger-Approximated Clonal Expansion Rate (PACER), which estimates mCA expansion rate from a single blood draw to build upon my measured mCA analysis by two orders of magnitude. A genome-wide association study and a phenome-wide association study will be conducted to identify germline variants and phenotypic correlations related to mCA clonal expansion rates. My working hypothesis for Aim 2 is that 1) certain germline variants predispose individuals to faster mCA growth and 2) specific disease phenotypes, including chronic lymphocytic leukemia and infection susceptibility, are associated with faster mCA clonal expansion rate. This research aims to significantly enhance our understanding of mCA clonal expansion, addressing a fundamental biological mechanism of aging to prevent multiple diseases. Collectively, these insights will contribute to mCA risk prediction models and highlight potential biological pathways or lifestyle strategies to slow mCA expansion.

NIAID - National Institute of Allergy and Infectious Diseases

Abstract CD8+ T cells are unique in effective sensing and killing of intracellular pathogen-infected cells and tumor cells. Because current vaccines are designed to induce high titer pathogen-specific antibodies for host protection, new vaccines focused on promoting effective memory CD8+ T cells are needed. Since a single naive CD8+ T cell has the potential to give rise to multiple types of progenies, it is essential to understand how naive T cells are primed to form distinct effector and memory cells. It is generally accepted that the strength of cognate antigen (Ag) stimulation determines the size of the primary response and of the memory cell pool, and that strong cognate Ag signals coupled with robust co-stimulation and cytokines altogether drive naive CD8+ T cells towards an effector rather than a memory cell fate. The current dogma also states that cognate Ag stimulation does not lead to functionally distinct subsets of memory CD8+ T cells. In contrast, however, we recently discovered that the strength and the stability of cognate Ag/MHC interactions with the T cell receptor (TCR) determine the development of memory cell functional characteristics, in particular stem-cell associated characteristics, through epigenetic imprinting. Stem cell memory CD8+ T (TSCM) cells have been shown to exhibit superior functional features, progeny potential, self-renewal capacity and longevity. Using state of the art conditional mouse models, high dimensional spectral flow cytometry, lentiviral-based inducible gain or loss of function experiments, and computational modeling approaches, we will define the features of T cell epitopes, key TCR structural modes of recognition, TCR signaling pathways, genetic and epigenetic regulators that enhance the differentiation of TSCM cells in vivo. We will validate our findings in models of chronic infections and tumors. This research directly impacts the rational design of more effective vaccines and adoptive T cell transfer therapies.

NCI - National Cancer Institute

Project Summary/Abstract Somatically acquired mutation in the E1 ubiquitin-activating enzyme UBA1 within hematopoietic stem and progenitor cells (HSPCs) was recently identified as the cause of the adult-onset autoinflammatory syndrome VEXAS (vacuoles, E1 enzyme, X linked, autoinflammatory, somatic). Most VEXAS-associated mutations occur at Met41 in UBA1 and lead to clonal expansion within the HSPC and myeloid compartments, and frequently cause myelodysplastic syndrome (MDS) in addition to multi-organ inflammation. Despite the severity and prevalence of VEXAS, the mechanisms whereby UBA1 mutations cause multiorgan autoinflammation and myeloid malignancy are unknown. Our preliminary work utilizes newly developed and genetically controlled primary cell-based models of VEXAS to show that UBA1 mutant HSCs are primed towards myeloid differentiation and that their myeloid progeny undergo aberrant inflammatory cell death upon engagement of innate immune receptors. We hypothesize that aberrant inflammatory cell death underlies VEXAS pathogenesis. To build upon our preliminary work, this career development program will address 2 specific aims: (1) to determine cell-intrinsic and -extrinsic factors that promote clonal expansion and myeloid skew of UBA1 mutant HSPCs, and (2) to evaluate the role of inflammatory cell death as a therapeutic target in VEXAS. The proposed studies are part of the candidate’s developing research program at the intersection of myeloid malignancy and innate immune signaling and build upon his clinical expertise in MDS and AML. Dr. Narendra will develop a research program under the close supervision of his mentor Dr. Alexander Gitlin, an expert in the molecular basis of innate immune signaling, and co-mentor Dr. Scott Lowe, an expert in cancer biology, mouse models and functional genetics. Dr. Narendra will additionally be supported by members of his advisory committee including Drs. Omar Abdel- Wahab, Caleb Lareau, and Alexander Rudensky, experts in experimental and computational approaches to the study of hematopoiesis and immunity. Under the guidance of his mentors and advisory committee, the candidate will continue to gain expertise in immunology, malignant hematopoiesis, and single cell analytics. Dr. Narendra’s training environment will ideally position him to achieve his goal of becoming an independent physician-scientist, working at the convergence of innate immunity and cancer.

NIAID - National Institute of Allergy and Infectious Diseases

Abstract. In the last 10 years, the field of induced pluripotent stem cells (iPSCs) has come far, but the reprogramming process is still sub-inefficient; around 1-3% of the transduced cells reprogram, and the iPSCs are just starting to be translated into a clinical trial. Therefore, there is a need for an efficient reprogramming system that would allow the in-depth analysis of the different phases of the reprogramming process and the role of the innate immune response. We have developed a new vector system, based on a human Paramyxovirus, measles virus (MeV). From this virus, we have shown that the measles virus can be modified into a “one cycle” reprogramming vector expressing the four reprogramming factors (RFs, OCT4, KLF4, SOX2 and cMYC) in one single genome and successfully generate genomic modification-free iPSCs from human fibroblasts. We showed that the MeV vector is equivalent to or superior to the Sendai (SeV) vector and that the MeV-derived iPSC clones present all the characteristics of iPSC clones. The long-term goal is to understand the process of reprogramming by MeV vectors to produce better human iPSCs for the treatment of degenerative diseases. The central hypothesis is that because the MeV reprogramming vector expresses the four reprogramming factors in one single vector, it can be used as a tool to understand the underlying process of iPSC reprogramming. The objectives of this particular application are to (1) identify the difference in the state of the innate immune response activation during MeV vector reprogramming, (2) understand how the control of the innate immunity by MeV affects reprogramming efficiency, and (3) understand the role of exogenous RFs expression in the reprogramming process. The proposed work is innovative because it capitalizes on a new MeV vector expressing the four RFs for the reprogramming of somatic cells into iPSC, and our group developed this technology. Also, the RFs expression can be further modulated in an innovative way by either post-transcriptional regulation, using miRNA that are naturally up-regulated or down-regulated during the reprogramming process or by changing their position in the genome. The proposed work is significant because it will collectively validate MV vector as a platform to produce clinically relevant iPSCs, but also as a new tool to study the reprogramming process. This work will lead to the identification of pathways important in MeV reprogramming, but these pathways could be investigated in the context of the related SeV vector and other reprogramming technologies.

NCI - National Cancer Institute

SUMMARY Basal-like breast cancer (BLBC) is an aggressive cancer subtype that disproportionally affects younger women and women of African ancestry, contributing to cancer disparity. Additionally, women with BRCA1 germline mutations have an extremely high risk of developing BLBC. BLBC precancer tends to be high-grade and likely to progress to malignant cancer. Understanding immune escape mechanisms in BLBC precancer is crucial for developing immunoprevention strategies to intercept its progression to aggressive breast cancer. Recent studies have showed that the generation of embryonic multipotent stem-like cells (referred to as pre-CSCs) through de- differentiation is critical for BLBC precancer progression in mouse models. Importantly, de-differentiation to multipotent cells has also been observed in patients with germline BRCA1 mutations. Similar de-differentiation processes occur in other breast cancer subtypes, suggesting a broad strategy for cancer prevention by targeting pre-CSCs. However, how pre-CSCs escape immunosurveillance remains poorly understood. In our preliminary studies, we have discovered that BLBC pre-CSCs express high levels of the stem cell transcription factor SOX9, which greatly induce the expression of the immune checkpoint B7x (also known as B7-H4, B7S1 or VTCN1). These cells also upregulate multiple cancer testis antigens. We found that the SOX9- B7x pathway is necessary for inhibiting T cell infiltration and protecting pre-CSCs from T cell-mediated elimination. SOX9 or B7x knockout blocks the progression of BLBC precancer. These findings strongly support the hypothesis that BLBC pre-CSCs are unexpectedly immunogenic due to upregulation of cancer testis antigens, and the upregulation of immune checkpoint B7x by stem cell factor SOX9 in these cells are required for establishing an immuno-suppressive microenvironment crucial for safeguarding pre-CSCs from immune elimination. This hypothesis will be tested by pursuing the following specific aims: (1) Dissect the mechanisms by which SOX9 regulates B7x and other immunosuppressive pathways in in precancer stem cells; (2) Determine the mechanistic interactions between precancer stem cells and their reactive immune cells; and (3) Develop anti- B7x therapy, either alone or in combination, as new immunoprevention strategies targeting pre-cancer stem cells in breast cancer. The most effective strategies will be further tested in mouse models of other breast cancer subtypes. To accomplish our goals, we have generated a number of novel tools and resources. The outcomes of this project will elucidate new immunosuppressive mechanisms of pre-CSCs and develop anti-B7x therapy, either alone or in combination, as immunoprevention approaches against breast cancer.

NCI - National Cancer Institute

This application is focused on the study of bioengineered plant virus-based adjuvant and vaccine technology. We discovered that some plant viruses serve as potent adjuvants in the context of infectious disease and cancer vaccines/immunotherapy. Cowpea mosaic virus (CPMV) was identified as a uniquely potent adjuvant with distinct mechanism of immunomodulation compared to small molecule agonists, other plant viruses, or oncolytic viruses. Recently we discovered that systemic CPMV administration prior to tumor challenge protects mice from onset of tumor growth. Data indicate that the innate immune stimulation by CPMV is durable and lasts for weeks after CPMV exposure when innate cells would have returned to a homeostasis state – therefore data are consistent with induction of trained immunity. Single cell sequencing analysis of human PBMCs after CPMV adjuvant exposure indicates stimulation of interferon signaling pathways along with metabolic changes, and epigenetic rewiring – also consistent with a mechanism involving trained immunity. Together our data suggest that CPMV could act as an inducer of trained immunity – to date there are no reports on the study of plant viruses in trained immunity. Proposed studies will help elucidate the foundational principles that make CPMV a uniquely potent immunomodulator. We will fulfil the following specific aims: (1) We will establish the mechanism of CPMV as a training agent in vitro using immune cells followed by LPS challenge; longitudinal studies will be carried out and analysis will include measure of pro- inflammatory cytokine as well as CHiP and ATAC sequencing to confirm epigenetic rewiring and metabolic cell changes. Structure-function studies of bioengineered viruses will provide foundational insights into differential potency. (2) We will establish the mechanism of CPMV as a training agent using the B16F10 tumor model using WT and Rag 1 vs CCR2 knockout (KO) mice to delineate the role of adaptative vs. innate immune cells (β-glucan will serve as benchmark). Hematopoietic stem cells (HSCs) and multipotent progenitors (MPPs) will be analyzed by single cell sequencing, CHiP and ATAC sequencing to delineate the mechanism of action. Studies will be paralleled with safety and biodistribution studies. (3) We will test the ability of the CPMV training agent to facilitate protection from influenza virus challenge; protection from influenza virus infection and pathology from CPMV will be benchmarked against FLUMIST and β-glucan. These studies could lay the foundation for continued and deeper studies of CPMV as an adjuvant technology towards the development of more broadly protective and efficacious vaccine formulations and immunoprevention strategies.

NINDS - National Institute of Neurological Disorders and Stroke

PROJECT SUMMARY BCL11A is a zinc-finger transcription factor that has been well-studied in erythroid biology, but evidence for an important role in the brain is beginning to emerge. Patients with heterozygous loss-of-function BCL11A mutations present with clinical features that can include intellectual disability (ID), autism spectrum disorder (ASD), and epilepsy. BCL11A is identified as a high confidence ASD risk gene in the SFARI database, and multiple lines of evidence also support potential roles in the etiology of schizophrenia (SZ) and Alzheimer’s disease (AD). However, the mechanisms that link BCL11A to these clinically challenging disorders are poorly understood. To better understand the neuronal function of BCL11A, we performed a series of preliminary behavioral analyses following the selective heterozygous deletion of Bcl11a from excitatory and/or inhibitory neurons in the mouse brain. Surprisingly, we found that Bcl11a deletion from inhibitory GABAergic interneurons (GINs) resulted in social deficits, hyperactivity, and increased seizure susceptibility. Furthermore, we observed increasing levels of BCL11A expression and physical occupation at predicted binding motifs during differentiation and maturation of GINs derived from human induced pluripotent stem cells (iPSCs). Additionally, we found that GIN-enriched ventral forebrain organoids derived from BCL11A-null iPSCs display differential gene expression signatures that overlap with pathological changes in the prefrontal cortex of postmortem brains of individuals with ASD and SZ. Taking these observations together, we hypothesize that the clinically challenging neurological phenotypes associated with BCL11A mutations likely reflect the specific impact of altered BCL11A function on different classes of neurons, with GINs being particularly vulnerable. We will test this hypothesis through a comprehensive series of in vitro (Aim 1) and in vivo (Aim 2) approaches. In Aim 1, we will identify and compare the gene targets of BCL11A in human iPSC-derived excitatory neurons and GINs. We will also employ single-nucleus (sn)RNA-seq and snATAC-seq along with whole-cell patch clamp electrophysiology and histology to establish the overlapping and distinct roles of BCL11A in human neuron populations. In Aim 2, we will further explore the in vivo function of Bcl11a by determining the behavioral and physiological effects of deleting Bcl11a in a neuron type-specific manner in mice. We will also use a chemogenetic approach to further interrogate GIN subtype-specific contributions to BCL11A disease mechanisms. Our long-term goal is to translate these findings into a better understanding of the role of BCL11A in the brain, which will help guide treatment development for patients with BCL11A dysfunction and other GIN-associated disorders.

NIDDK - National Institute of Diabetes and Digestive and Kidney Diseases

Abstract Intestinal stem cells (ISCs) play pivotal roles in intestinal epithelium renewal during homeostasis and after injury. The metabolic demands faced by ISCs require high mitochondrial oxidative phosphorylation (OXPHOS) activity compared to other differentiated cells. ISCs’ mitochondrial dysfunction has been implicated in the etiopathogenesis of intestinal bowel diseases (IBD), which afflicts over 2 million people in the US. The carbon sources that fuel ISC OXPHOS have been broadly described in the small intestine (SI) but not in the colon. I seek to understand how ISC metabolic demands are met in the colon. The colonic epithelium is organized into the colonic crypt. ISCs localize to the base of the crypt (base-crypt), and this protects them from microbial metabolites and microbe-associated molecular patterns (MAMPs). Top-crypt differentiated colonic epithelial cells (CECs) oxidize microbial-derived short-chain fatty acid (SCFA) butyrate making it inaccessible to base-crypt cells. This shields ISCs, as butyrate suppresses ISC proliferation. This metabolic interaction between CECs and ISCs has focused my interest in CEC-ISC metabolic cross-talk. Ketones (acetoacetate, β-hydroxybutyric acid (βHB), and acetone) are important metabolic substrates. I hypothesize that CECs generate ketones that are used by ISCs as their principal energy source. This hypothesis is supported by the localization of rate-limiting enzymes (RLE) for the generation of ketones to the CECs and my preliminary data demonstrating that loss of these enzymes in CECs compromises ISC self-renewal and differentiation. My proposal focuses on this metabolic cooperation between epithelial cells within the crypt where top-crypt CECs shuttle ketones to base-crypt ISCs thus maintaining their turnover capacity. Understanding colonic ketone biosynthesis and function could lead to new treatments and therapeutic targets for IBD. Beyond defining this proposed metabolic crosstalk between CEC and ISC, I am interested in factors regulating CEC metabolic enzymes. I have identified microbial features that regulate the expression of the CEC RLE for ketone generation. I will determine the receptor and pathways downstream of this receptor by which they regulate the expression of this RLE. This research will help to decipher how microbial signals and metabolites contribute to epithelial repair and regeneration.

NIGMS - National Institute of General Medical Sciences

PROJECT SUMMARY Stem cells and progenitors are integral for tissue development, homeostasis and regeneration, and their dysfunction underlie development disorders, defective tissue regeneration, aging and cancer. In addition to intrinsic mechanisms, diverse niche cells influence stem/progenitor cell fate. Adult tissues harbor heterogenous stem/progenitor cell subsets which are intricately involved in tissue regeneration and disease. Yet, the fundamental cell types and molecular mechanisms that regulate distinct tissue-specific stem/progenitor cell populations remain poorly understood. Using the murine mammary gland as a model, we were the first to uncover primitive mesenchymal progenitor niche cells that have a remarkable capacity to contribute to a major subset of tissue-specific epithelial progenitors, indicating their potential role in replenishing progenitor cells central to tissue regeneration. We have also observed that sympathetic neurons modulate progenitor subsets in the niche and epithelium to orchestrate tissue regeneration. These niche cell types permeate diverse somatic tissues and thus, understanding their control of tissue-specific progenitors is critical for advancing progenitor- targeted therapies in regenerative medicine and malignancy. The proposed research will investigate how tissue-specific progenitor cell fate is dictated by mesenchymal progenitors and sympathetic neurons during adult tissue regeneration. We will utilize the postnatal mouse mammary gland as a tractable model to study tissue-specific progenitors given that it undergoes extensive morphogenesis directed by progenitors. We will explore niche cell-mediated modulation of progenitors and identify underlying molecular signals and mechanisms. We will employ a variety of strategies that include in vivo genetic reporter, lineage tracing and cell ablation mouse models and ex vivo organoids combined with phenotypic, functional assays and next-generation sequencing. Our findings will bridge key gaps in knowledge of the niche-driven circuitry that impacts tissue progenitors and provide a framework for improving the outcome of diseases attributed to aberrant progenitors.

U.S. National Science Foundation

This program will advance scientific community datasets to enable scientific discovery and innovation using artificial intelligence (AI) and other methods. Its goal is to increase the value that can be derived from existing scientific datasets by leveraging novel methods and artificial intelligence (AI). This will unlock new AI-driven insights. It can enable interdisciplinary research. It also enables investigations outside of the original motivation for data collection and analysis.The program seeks to 1) apply AI-based capabilities to feature extraction and metadata generation, and the integration of multiple datasets. 2) Develop robust data pipelines necessary for automated analysis of existing datasets and similar use-cases by AI tools and systems. 3) Augment and/or harmonize existing datasets to better enable use by AI data pipelines and automated analysis. Proposals should address dataset security and integrity. Governance and the process for scientific communities to contribute to the datasets should also be addressed. Proposals are encouraged to leverage existing resources. These may include NSF data platforms, the NSF Integrated Data Systems and Services program, the NSF-led National AI Research Resource, the Genesis Mission platform, or other national infrastructure.NSF is open to exploring partnerships with philanthropy, private industry, or the non-profit sector to support additional proposals or collaborative opportunities that will advance AI-driven scientific discovery through unlocking the value of high-impact scientific datasets.Expanding Participation in STEM, NSF Priorities, and Gold Standard Science:NSF prioritizes cutting-edge discovery science and engineering research, advancing technology and innovation, and creating opportunities for all Americans. NSF has established priorities set forth by Congress, the administration and the NSF director to promote NSF's mission. Proposers should review the list of NSF priorities and are encouraged to align their proposals with them, where appropriate. NSF also expects the highest standards of scientific rigor, integrity and adherence tenets of Gold Standard Science in proposals, as appropriate for the field of science and research modality.