Proteomic Identification of the MYST Domain Histone Acetyltransferase TIP60 as a Coactivator of the Myeloid Transcription Factor C/EBPα

The transcription factor C/EBPα is a key player in granulopoiesis
and leukemogenesis. In the present study, we sought to identify
C/EBPα interacting proteins. A glutathione-S-transferase-C/EBPα
fusion protein was used to pull down interacting proteins from U937
nuclear extracts. These proteins were analyzed by 2-D gel
electrophoresis or 1-D nano LC and identified by mass spectrometry.
The interaction between C/EBPand two novel interacting partners,
the cell cycle regulator protein MCM5 and the MYST domain histone
aceyltransferase TIP60, was confirmed by using pull-down and
co-immunoprecipitation experiments. TIP60 was able to markedly
enhance C/EBPα mediated transcriptional activation in reporter gene
assays, suggesting that TIP60 is a co-activator of C/EBPα. This
co-activator function of TIP60 was dependent on its intact histone
aceyltransferase domain and on the C/EBPα DNA binding domain. TIP60
was found to be associated with the human C/EBPα promoter in vivo
in a chromatin immunoprecipitation assay with a concomitant
increase in histone H3 and H4 acetylation. Furthermore, we observed
a lower expression of TIP60 mRNA in undifferentiated U937 CD11b-
cells compared to retinoic acid induced differentiated U937 CD11b+
cells suggesting that higher TIP60 expression is associated with
myeloid differentiation. Correlated expression between C/EBP and
TIP60 was also observed in certain leukemia subtypes. These
findings point to a functional synergism between C/EBP and TIP60
in myeloid differentiation and suggests that TIP60 might be an
important player in leukemogenesis.