Investigating the developmental and transcriptional bases for distinct functions of IL-10+ and IL-10− Treg cells - PROJECT SUMMARY/ABSTRACT The finely tuned generation and function of regulatory T (Treg) cells are essential for maintaining the balance that allows for protective immunity while preventing harmful autoimmunity. Treg cells are heterogeneous, comprising specialized subsets that contribute to tissue repair and mediate context-specific immune responses. Despite their critical roles in essential biological processes, it remains unknown whether the subset-specific functions of Treg cells are driven by their developmental origins, transcriptional programs, or a combination of both. This unresolved challenge largely stems from two issues: the lack of unbiased means to identify mutually exclusive Treg cell subsets with distinct functions, and the absence of tools to trace their ontogeny. However, my recent discoveries have opened promising avenues for overcoming these obstacles. Using colorectal cancer models and human patient specimens, I identified that interleukin-10 (Il10) expression distinguishes two subsets of Treg cells with opposing functions: IL-10+ Treg cells, which exhibit anti-tumor properties, and IL-10– Treg cells, which promote tumor growth. Furthermore, I identified Dapl1 as a gene uniquely expressed by naïve CD4 T cells, thereby providing a definitive marker for extrathymically generated Treg cells. The overarching goal of this research proposal is to determine whether the developmental origins of IL-10– vs IL-10+ Treg cells contribute to their distinct functions, and to identify the transcriptional programs underlying these differences. This proposal tests the hypothesis that both of these subsets are of mixed developmental origins, with their distinct functions driven by differentially expressed transcription factors. Specifically, in Aim 1, using a novel Dapl1-based lineage tracing model, I will determine whether IL-10+ and IL-10– Treg cells arise from thymic or extrathymic (peripheral) origins and elucidate how these developmental pathways shape their functions. Additionally, in Aim 2, I will define the transcriptional programs that drive their subset-specific activities, by inducing Treg cell specific deletion of key regulators such as Zeb2 and Nfil3. By employing genetic mouse models, advanced single-cell analyses, and CRISPR-based screening, the proposed studies will reveal the nature of Treg cell functional heterogeneity, ultimately guiding the development of more precise immunotherapeutic strategies with major implications for public health. The proposed career development plan complements my training in cellular and molecular immunology with single-cell analysis and computational biology. I will take advantage of the extensive resources of the Memorial Sloan Kettering Cancer Center, part of the Tri-Institutional network with the Rockefeller University and Weill Cornell, as well as benefit from the mentorship of Dr. Alexander Rudensky and guidance from Advisory Committee members Dr. Christina Leslie, Dr. Ming Li, and Dr. Steven Josefowicz. By the end of the mentored phase, I will have acquired the necessary tools to conduct comprehensive studies at the intersection of immune cell heterogeneity and immune communication with the environment as an independent investigator.
