Cyp33-MLL Interaction Modulates Human HOX Gene Expression and Chromatin Structure.

The MLL gene is involved in translocations and fusion genes associated with human leukemia. We have identified a protein, Cyp33, which interacts with the PHD fingers of MLL, and modulates the expression of HOX genes regulated by MLL. In order to define possible mechanisms for this modulation, we set up an experimental system in which the Cyp33 protein is conditionally expressed from a transgene under the control of a tetracycline inducible promoter. Two different cell lines with such inducible gene were established, based on the human kidney epithelial cells HEK293, and the human lymphoblastic leukemia cell line Jurkat, respectively. In these systems we have shown that induction of Cyp33 expression over endogenous levels, leads to downregulation of the HOXC8 gene, but this downregulation is reversible, the HOXC8 being re-expressed when Cyp33 is de-induced and its levels go back to normal. This indicates that Cyp33 overexpression does not erase an epigenetic mark for HOXC8 gene expression. Concordantly, no change in methylation of histone H3 at lysines 4 or 9 is observed upon Cyp33 overexpression. We also showed that both MLL and Cyp33 bind to the promoter of HOXC8, and that histones H3 and H4 at this promoter are de-acetylated upon Cyp33 overexpression. This deacetylation, as well as the downregulation of HOXC8 expression, are prevented by cyclosporin A and by trichostatin A, suggesting that the prolyl-isomerase activity of Cyp33 is necessary for these effects, and that histone deacetylation mediates these effects. We have previously shown that overexpression of Cyp33 enhances the recruitment of HDAC1 to the repression domain of MLL. We now show that this enhanced recruitment is inhibited by cyclosporin A. These results suggest a model in which Cyp33 is recruited to MLL and targets the MLL repression domain to promote the recruitment of histone deacetylases, thus allowing or promoting the silencing of the MLL regulated genes. In this model Cyp33 functions downstream of the gene activation or silencing epigenetic marks.