The leukaemogenic fusion protein MOZ-TIF2 inhibits nuclear receptor-mediated transcription and mislocalises CBP
Chromosomal rearrangements associated with the M4/M5 subtype of Acute Myeloid Leukaemia include fusions of the gene encoding the histone acetyltransferase (HAT) MOZ with genes encoding the transcription factor coactivators TIF2, CBP or p300. The physiological roles of MOZ in vivo remain to be established. However, TIF2 functions to recruit the HATs CBP/p300 to ligand-bound Nuclear Receptors (NR), thus leading to transcription activation. Studies investigating the role of NRs and their ligands in haematopoiesis suggest that they aid the regulation of myeloid cell differentiation. The research described in this thesis therefore investigated the effect of the MOZ-TIF2 fusion protein on NR-mediated transcription. Expression of MOZ-TIF2 resulted in an inhibition of ligand-dependent transcription by the Retinoid X Receptor a, Retinoic Acid Receptor a and Estrogen Receptor a in the C0S1 cell line. Further studies showed that MOZ-TIF2 also inhibited transcription mediated by endogenous NRs in the haematopoietic cell line U937. Investigation of protein interactions demonstrated that MOZ-TIF2 bound CBP through its Activation Domain 1 (AD1) and that deletion of this region resulted in the inability of the protein to inhibit NR transcription. Immunofluorescence analysis showed that MOZ-TIF2 formed a mesh-like nuclear pattern in contrast to the speckled nuclear localisations of MOZ, TIF2 and CBP. In addition, in cells expressing MOZ-TIF2, but not the AD1 deletion mutant, endogenous CBP displayed a diffuse staining pattern. Thus, the experiments described here show that MOZ-TIF2 inhibits NR-mediated transcription and mislocalises CBP, and that both of these activities are dependent upon its AD1. Further experiments investigating the effect of MOZ-TIF2 on p53 and AML1 -mediated transcription have indicated that MOZ-TIF2 also has the ability to affect the function of transcription factors in addition to NRs. Thus, expression of the MOZ-TIF2 fusion protein may result in the formation of leukaemia through the misregulation of normal haematopoietic transcription.