DOT1L elicits mitotically heritable changes to gene expression through the methylation of lysine 79 on Histone 3 and is a major driver of malignant gene expression in MLL-rearranged B-cell Acute Lymphoblastic Leukemia (MLLr B-ALL); an aggressive cancer with a particularly poor prognosis in the children and adults affected by it. Consequentially, phase I clinical trials have begun testing DOT1L-inhibitors in MLLr-ALL patients. Despite this, the role of DOT1L in normal B-cell biology remains largely unknown. Herein, we sought to characterise the role of DOT1L in the regulation of the humoral immune response.
Dot1lf/fCd23Cre/+ mice were immunised with a range of T-dependent and T-independent antigens and their B-cell responses were compared against wildtype controls. The conditional deletion of Dot1l resulted in a significant attenuation of early memory B-cells, class-switched PCs and GCs produced in response to the antigen. RNA-seq revealed that genes associated with B-cell migration and signalling, such as S1pr2, Ackr2, Ackr4, Tnfrsf17 and Gcsam were upregulated upon activation in control B-cells, but not in Dot1l deficient B-cells. Migration assays and histological analysis revealed that Dot1l-deficient activated B-cells failed to migrate into the B-cell follicle to form GCs, and instead accumulated in follicular borders. H3K79me2 ChIP-seq was performed on naive and GC B-cells and confirmed that the majority of the genes downregulated in Dot1l-deficient activated B-cells were direct gene targets of DOT1L. Moreover, DOT1L was found to be targeting genes associated with biological processes that are central to the operation and maintenance of a fully formed GC, such as V(D)J recombination, DNA repair and cell division. Together, these results reveal a vital role for DOT1L in regulating various stages of B-cell differentiation and in the generation of effective and lasting humoral immunity.