Dependence on Myb expression is attenuated in myeloid leukemia with N-terminal CEBPA mutations

Mutations altering the normal function of C/EBPα are frequent in acute myeloid leukaemia with normal karyotype. MYB, a cooperating partner of C/EBPα, is likewise heavily implicated in AML. Here we investigate how the relative requirement for the transcription factor MYB in AML relates to the particular combinations of wild type and mutated alleles of CEBPA. Through knockdown of Myb in murine cell lines modelling the spectrum of CEBPA mutations we show that the consequences of reduced Myb depend on the mutational status of Cebpa. Importantly, Myb knockdown fails to override the block in myeloid differentiation in cells with biallelic N-terminal C/EBPα mutations, demonstrating for the first time that the dependency on Myb observed in AML is much lower in leukaemia with this combination of mutations. By comparing genome-wide analyses of gene expression following Myb knockdown and ChIP-seq data for the binding of C/EBPα isoforms, we provide evidence for a functional cooperation between C/EBPα and Myb in the maintenance of the leukaemia state. This co-dependency breaks down when both alleles of CEBPA harbour N-terminal mutations, as a subset of C/EBPα-regulated genes only bind the short p30 C/EBPα isoform and, unlike other C/EBPα regulated genes, do so without a requirement for Myb. Gene expression analysis of FMH9, KL and LL cells with and without Myb knockdown