The Role of the YEATS Epigenetic Reader in MLL-r Fusion Proteins

Abstract

MLL (KMT2a) translocations are found in ~10% of acute leukemia patients, giving rise to an aggressive subset of leukemias. The majority of MLL fusion partners are members of epigenetic and transcriptional regulatory complexes, such as the Super Elongation Complex (SEC) and the DOT1L Complex (DotComm). In MLL-rearranged (MLL-r) leukemias, these protein complexes are hijacked to deregulate transcription and the epigenetic landscape of pro-leukemic genes. Thus, identifying druggable targets within these transcriptional and epigenetic complexes has become an attractive area of research for developing future therapeutics. Two common MLL translocation partners, ENL and AF9, are homologous proteins of the YEATS domain family. Both proteins contain a highly conserved N-terminal YEATS epigenetic reader domain that binds to histone acylation and interacts with the PAF1c, an epigenetic regulator protein complex essential for MLL-fusion leukemia. Recent work demonstrated the importance of wild-type ENL, and specifically its YEATS domain, in acute myeloid leukemia (AML) growth. In contrast, AF9 is dispensable for AML proliferation. This pointed to the ENL YEATS domain as a potential AML therapeutic target but did not address whether the YEATS domain impacts MLL-ENL fusion protein oncogenesis. Thus, we sought to investigate the YEATS epigenetic reader domain in the context of the MLL-ENL fusion proteins to uncover potential therapeutic opportunities. In my thesis work, we found that the YEATS domain is retained in the majority (84.1%) of MLL-ENL fusions in t(11;19)(q23;p13.3) patients (N=302) and excluded in almost all MLL-AF9 fusions in t(9;11)(p22;q23) patients (N=449). This striking difference in YEATS domain inclusion between the MLL-ENL and MLL-AF9 fusion proteins prompted us to investigate the significance of the YEATS domain on MLL-ENL fusion proteins. Through our biochemical and biological studies, we show that the ENL YEATS epigenetic reader function critically modulates MLL-ENL fusion protein mediated leukemic stem cell (LSC) frequency. Indeed, global transcriptomic analyses confirmed that the YEATS domain impacts MLL-ENL target genes involved in LSCs. Genetic perturbation of the MLL-ENL YEATS epigenetic reader function significantly extends leukemia latency, while absence of the YEATS domain and downstream sequence abrogated leukemogenesis in vivo. Mechanistically, YEATS domain deletion impaired the MLL-ENL fusion protein and PAF1c binding at a subset of MLL-ENL targets. Mutations in the MLL-ENL YEATS reader pocket led to depletion of active histone marks and compromised expression of a subset of MLL-ENL targets, including the transcriptional factor Eya1. Therapeutically, the YEATS domain sensitizes MLL-ENL fusion driven leukemic cells to YEATS inhibitor treatment. Our results demonstrated an oncogenic role for an epigenetic reader presented on a subset of MLL-r fusion proteins. We functionally linked the YEATS epigenetic reader with leukemic stem cell (LSC) frequency in MLL-ENL leukemias and showed its values as a therapeutic target. Potentially, YEATS inhibitor could be utilized either as a single agent or in part of a combination therapy in treating MLL-ENL leukemia and/or drug-resistant AML. Our data further contributes to the greater understanding of ENL versus AF9, in how different parts of these proteins (i.e., the YEATS domain, the internally disordered region (IDR), the ANC-1 homology domain (AHD)) contribute to their roles as key epigenetic and transcriptional regulators in normal and/or malignant hematopoiesis.