ABSTRACT
The mixed lineage leukemia (MLL) gene plays a critical role in epigenetic regulation of gene expression and is a frequent target of chromosomal translocations leading to leukemia. MLL plant homeodomain 3 (PHD3) is lost in all MLL translocation products, and reinsertion of PHD3 into MLL fusion proteins abrogates their transforming activity. PHD3 has been shown to interact with the RNA-recognition motif (RRM) domain of human nuclear Cyclophilin33 (CYP33). Here, we show that CYP33 mediates downregulation of the expression of MLL target genes HOXC8, HOXA9, CDKN1B, and C-MYC, in a proline isomerase-dependent manner. This downregulation correlates with the reduction of trimethylated lysine 4 of histone H3 (H3K4me3) and histone H3 acetylation. We have structurally characterized both the PHD3 and CYP33 RRM domains and analyzed their binding to one another. The PHD3 domain binds H3K4me3 (preferentially) and the CYP33 RRM domain at distinct sites. Our binding data show that binding of H3K4me3 to PHD3 and binding of the CYP33 RRM domain to PHD3 are mutually inhibitory, implying that PHD3 is a molecular switch for the transition between activation and repression of target genes. To explore the possible mechanism of CYP33/PHD3-mediated repression, we have analyzed the CYP33 proline isomerase activity on various H3 and H4 peptides and shown selectivity for two sites in H3. Our results provide a possible mechanism for the MLL PHD3 domain to act as a switch between activation and repression.
Subject(s)
Cyclophilins/physiology , Gene Expression Regulation , Myeloid-Lymphoid Leukemia Protein/physiology , Binding Sites , Down-Regulation , Histones/metabolism , Humans , Myeloid-Lymphoid Leukemia Protein/chemistry , Peptidylprolyl Isomerase/metabolism , Protein Binding , Protein Structure, Tertiary , Repressor Proteins , Up-RegulationABSTRACT
Retinoblastoma can arise due to mutational inactivation or methylation of RB1 gene promoter. A 600-bp CpG island consisting of the essential promoter is present at the 5' end of RB1 gene. Hypermethylation of the CpG island within the RB1 promoter region has been described in unilateral retinoblastoma. In vitro and in vivo studies have suggested that methylation of the RB1 promoter dramatically reduces gene activity. In the present study methylation status of the CpG island within the promoter region of RB1 gene has been evaluated by methylation specific polymerase chain reaction to define the molecular mechanism responsible for retinoblastoma in Indian patients. One unilateral and two bilateral nonhereditary patients had methylation of the RB1 promoter region in which 6.6% of our patients had complete methylation of the RB1 promoter region. This study shows methylation of RB1 promoter is not a major mechanism for retinoblastoma patients in India. Methylation analysis is used in genetic counseling of the family.
