Lymphocytes and their precursors are licensed to transiently activate specific mutation pathways that enable efficient remodeling of antigen-receptor genes. To generate the enormous diversity of clonotypic antigen receptors, specific DNA lesions are generated and resolved in an error-prone fashion at defined stages of lymphocyte development. These characteristics provide ideal model systems to study not only the role of DNA damage response (DDR) and DNA damage tolerance (DDT) pathways in resolving specific DNA lesions to shape the immunoglobulin (Ig) repertoire, but also in genome stability, stem cell maintenance, and tissue homeostasis.
Our research activities are focused on two subjects:
(i) DNA damage tolerance (DDT) in physiology and precision cancer medicine
(ii) Epigenetic programing of lymphocyte development and differentiation: From fundamental insights to translational research
Stem cells are key players in central biological processes, such as tissue homeostasis, ageing, and cancer formation. Stem cell depend on genome maintenance to prevent disease formation. DNA damage tolerance (DDT) pathways enable DNA replication in the presence of replication impediments and are regulated by PCNAK164 ubiquitination and REV1. The failure to generate PcnaK164R/K164R;Rev1-/- deficient mice revealed DDT as essential for mammalian life. The compound mutation rendered hematopoietic stem cells (HSCs) and the hematopoietic precursors genetically unstable, instigating a pathological process where the associated HSC depletion culminated in a severe, embryonic-lethal anemia. Single cell RNA-sequencing of the remaining HSCs and progenitors identified CD24Ahigh and CD93low erythroid-biased progenitors (EBP) within the Lineage-, Sca1+, cKit- (LSK) population. In line, this subset was found to depend on the erythroid transcription factor Klf1. In conclusion, DDT is an essential activity within the DNA damage response network and in maintaining HSC fitness. By studying this system, we identified an erythroid-biased progenitor subset within the LSK compartment.
Differentiation of naïve peripheral B cells into terminally differentiated plasma cells is characterized by epigenetic alterations, yet the epigenetic mechanisms that control B cell fate remain unclear. Here we identified a central role for the histone H3K79 methyltransferase DOT1L in controlling B cell differentiation. Naïve and activated murine B cells lacking Dot1L prematurely acquired plasma cell features and failed to establish germinal centers (GC) and normal humoral immune responses in vivo. Mechanistically, combined epigenomics and transcriptomics analysis revealed that DOT1L promotes expression of a pro-proliferative (Myc) and pro-GC program (Bach2) and supports the expression of the H3K27 methyltransferase Ezh2, the catalytic component of Polycomb Repressor Complex 2 (PRC2). Thereby, DOT1L ensures PRC2-mediated repression of anti-proliferative and plasma cell differentiation program. Our findings show that DOT1L is a critical regulator of the core transcriptional and epigenetic landscape in B cells and establishes an epigenetic barrier warranting B cell naivety. Details, see bioRxiv: doi.org/10.1101/826370).
Similar observations were made in the CD8+ T cell lineage.
Cytotoxic T-cell differentiation is guided by epigenome adaptations, but how epigenetic mechanisms control lymphocyte development has not been well defined. Our data indicate, that the histone methyltransferase DOT1L, which marks the nucleosome core on active genes, safeguards normal differentiation of CD8+ T cells. T-cell specific ablation of Dot1L resulted in loss of naïve CD8+ T cells and premature differentiation towards a memory-like state, independent of antigen exposure and in a cell-intrinsic manner. Mechanistically, DOT1L controlled CD8+ T-cell differentiation by ensuring normal T-cell receptor density and signaling. DOT1L also maintained epigenetic identity, in part by indirectly supporting the repression of developmentally-regulated genes. Finally, deletion of Dot1L in T cells resulted in an impaired immune response. Through our study, DOT1L is emerging as a central player in physiology of CD8+ T cells, acting as a barrier to prevent premature differentiation and controlling epigenetic integrity. Our findings highlight the importance of epigenetic signals in regulating T cell differentiation. Understanding these epigenetic signals is important for developing strategies to modulate T cell immunity. PNAS 2020, in press.