Genetic Analysis of T-cell Differentiation

Division Of Basic Sciences - Nci

Investigators

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Abstract

We study T lymphocytes (T cells), white blood cells essential for immune responses. Most express an antigen receptor (TCR) comprising a dimeric alpha-beta antigen-binding subunit (ab T cells) and recognize peptide antigens presented by class I (MHC-I, HLA A, B, and C in humans) or class II (MHC-II, HLA DP, DQ and DR in humans) classical Major Histocompatibility Complex molecules. Most such T cells express either of two surface glycoproteins that contribute to antigen recognition: CD4, which binds MHC-II, or CD8, which binds MHC-I. Consistent with such binding properties, MHC I-specific T cells generally are CD4-CD8+ (CD8 T cells), whereas MHC II-specific T cells generally are CD4+CD8- (CD4 T cells). CD4 T cells are essential for life: CD4 T cell deficiency, whether innate or acquired, leads to recurrent or chronic infections and death. Studies of ab T cell development in the thymus have remained at the forefront of the laboratory activity in the past few years. To decipher the transcriptional circuits that control thymocyte development, we are mapping the role of transcription factors putatively involved in this process using single cell (sc) RNA sequencing to analyze gene expression, and sc ATAC sequencing for chromatin accessibility. Additional ongoing studies investigate the mechanisms underpinning the strict requirement for the transcription factor Thpok (encoded by Zbtb7b) during CD4 T cell development and most responses to infections and tumors. The recent identification of a mutation in the human gene encoding Thpok, ZBTB7B, resulting in impaired CD4 T cell development and immunodeficiency, highlights the importance of mechanistic studies of Thpok functions. We are using similar approaches to explore the intrathymic development of "unconventional" intraepithelial lymphocytes (IEL), a subset of T cells initially described, and particularly abundant, in the gut where they express homodimers of the CD8a molecule ("CD8aa" IEL). Although the function of these cells remains largely enigmatic, cells of similar thymic origin colonize other epithelial organs and are thought to contribute to immune responses to tumors. Current work on thymic IEL precursors (IELp) follows up on our recent demonstration that the zinc finger transcription factor LRF (encoded by Zbtb7a) is necessary for the proper development of these cells (Nie et al, 2022). IELp are "agonist-signaled" thymocytes that carry TCRs with high affinity for self-ligands; as a result, many of them are deleted in the thymus by antigen-induced cell death ("negative selection"). We are exploring the mechanisms that allow some of these precursors to escape negative selection and differentiate in mature IELp that leave the thymus and colonize epithelia. Current work focuses on two questions. First, we are using single-cell and tissue transcriptomics to define the developmental trajectories of IELp, and identify their intrathymic interactions. Second, we are investigating the mechanisms that control expression of Bim, a pro-apoptotic protein involved in the negative selection of agonist-signaled thymocytes. In addition, the laboratory contributed to a study that found IEL in organs other than the gut, including the mammary gland, in which they are important for the development of lactating functions (Corral et al., 2025). In addition to T cell development studies, ongoing work explores CD4 T cell responses to infection and tumor antigens, building on recent publications from the laboratory (Ciucci et al., 2019; Magen et al., 2019; Ciucci et al., 2022; Chopp et al., 2023). Collaborative studies, notably using multiplex immunofluorescence and single-cell and tissue transcriptomics, are currently examining how CD4 T cells interact with intratumoral myeloid cells. We found that CD4 T cells can reprogram monocyte-macrophage type cells to switch from pro- to anti-tumoral functions; we are currently working to identify the signals involved and the effector mechanisms they trigger in macrophages.