Periodic Reporting for period 4 - REGiREG (Regulating the immune regulators: targeting adaptive immune control)
Reporting period: 2019-08-01 to 2021-06-30
For examples, we performed epigenetic analysis with a tagmentation-based whole-genome bisulfite sequencing technology, which allowed us to analyze the DNA methylation status of very few Treg cells isolated from tissues. With this technology, we were able to study the epigenetic changes of DNA methylation with the highest possible resolution (single nucleotide and genome-wide). Similarities of the epigenetic landscape led to the identification of a common tissue Treg population, present in many organs and characterized by gain and loss of DNA methylation, including many TH2-specific sites. Thus, we could show that tissue Treg cells integrate different waves of epigenetic reprogramming which define their tissue-restricted specializations. The corresponding manuscript has been published in Nature Immunology (Delacher M et al. Genome-wide DNA-methylation landscape defines specialization of regulatory T cells in tissues. Nature Immunology, 2017;18(10):1160-1172).
We wanted to understand the development of these tissue-resident Treg cells. By using our novel Nfil3-Cre-eGFP reporter mouse line, single-cell RNA-sequencing and bulk ATAC-sequencing, we identified two precursor stages of these IL-33 receptor ST2-expressing nonlymphoid tissue Treg population in spleen and lymph nodes. Molecular profiling of these precursors revealed their sequence of differentiation. Global chromatin profiling of nonlymphoid tissue Treg cells and the two precursor stages revealed a stepwise acquirement of open chromatin accessibility and reprogramming towards the nonlymphoid-tissue Treg cell phenotype. Mechanistically, we identified and validated the basic leucine zipper transcription factor, ATF-like (Batf) as the driver of the molecular tissue program in the precursors. We were able to publish these findings in the journal Immunity (Delacher et al., Precursors for Nonlymphoid-Tissue Treg Cells Reside in Secondary Lymphoid Organs and Are Programmed by the Transcription Factor BATF. Immunity, 2020; 18;52(2):295-312).
We were also able to shed light on human tissue-resident Tregs cells. We identified features that characterize human Treg cells with tissue-repair function. Single-cell chromatin accessibility profiles of murine and human tissue Treg cells defined a conserved, microbiota-independent tissue-repair Treg signature with a prevailing footprint of the transcription factor BATF. This signature, combined with gene expression profiling and TCR fate mapping, identified a population of tissue-like Treg cells in human peripheral blood that expressed BATF, chemokine receptor CCR8 and HLA-DR. Human BATF+CCR8+ Treg cells from normal skin and adipose tissue shared features with nonlymphoid T follicular helper-like (Tfh-like) cells, and induction of a Tfh-like differentiation program in naive human Treg cells partially recapitulated tissue Treg regenerative characteristics, including wound healing potential. Human BATF+CCR8+ Treg cells from healthy tissue share features with tumor-resident Treg cells, highlighting the importance of understanding the context-specific functions of these cells. The corresponding manuscript has been published in the journal Immunity (Delacher et al., Single-cell chromatin accessibility landscape identifies tissue repair program in human regulatory T cells. Immunity, 2021; 13;54(4):702-720).
We were also able to find an inhibitor of tissue-Treg cell differentiation. We described the function of the transcription factor Rbpj in Treg cells. Rbpj-deficient Treg cells adopt open chromatin landscapes and gene expression profiles similar to tissue Treg cells, with a prevailing signature of the transcription factor Gata-3. This study suggests that Treg cells require Rbpj to specifically restrain TH2 responses, including their own excessive TH2-like differentiation potential, thereby, inhibiting the tissue Treg pathway. Published in the journal Nature Communications (Delacher et al., Rbpj expression in regulatory T cells is critical for restraining TH2 responses. Nature Communications, 2019; 8;10(1):1621).
Regulatory T cells express the forkhead box transcription factor Foxp3 as a lineage-defining protein. Negative regulators of Foxp3 expression are not well understood. In our study, we generated double-stranded DNA probes complementary to the Foxp3 promoter sequence and performed a pull-down with nuclear protein in vitro, followed by elution of bound proteins and quantitative mass spectrometry. Of the Foxp3-promoter-binding transcription factors identified with this approach, one was T cell factor 1 (TCF1). Our data implicate a role of TCF1 in suppressing Foxp3 expression in activated T cells (Delacher et al., Quantitative Proteomics Identifies TCF1 as a Negative Regulator of Foxp3 Expression in Conventional T Cells. iScience, 2020; 22;23(5):101127).