The mutation of transcription factor genes is a main cause for acute myeloid leukaemia. RUNX1/ETO, the product of the t(8;21) chromosomal translocation, subverts normal blood cell development by impairing myeloid differentiation. RUNX1/ETO knockdown alleviates this block, with a global reprogramming of transcription factor binding and initiation of myeloid differentiation. Co-depletion of the myeloid transcription factor C/EBPα with RUNX1/ETO suppressed this differentiation response. Furthermore, C/EBPα overexpression largely phenocopied the effect of RUNX1/ETO knockdown. Our data show that low levels of C/EBPα are critical to the maintenance of t(8;21) AML and that C/EBPα drives the response to RUNX1/ETO depletion. To examine how changes in transcription factor binding impact on the activity of cisregulatory elements we mapped genome wide promoter-distal-element interactions in a t(8;21) AML cell line, via Capture HiC, and found that hundreds of interactions were altered by RUNX1/ETO knockdown. Differentially interacting elements exhibited changes in C/EBPα binding and were enriched for the CTCF motif. Our results demonstrate that the presence or absence of RUNX1/ETO has a profound impact on the intra-nuclear organisation of t(8;21) AML cells, and indicate which transcription factors are driving these changes. This work provides a novel mechanism for the RUNX1/ETO mediated differentiation block in t(8;21) AML.