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Southall, S. M., Wong, P. S., Odho, Z., Roe, S. M. & Wilson, J. R. Structural basis for the requirement of additional factors for MLL1 SET domain activity and recognition of epigenetic marks. Mol. Cell 33, 181-191

Institute of Cancer Research, Chester Beatty Laboratories, Chelsea, London, UK.
Molecular cell (Impact Factor: 14.46). 02/2009; 33(2):181-91. DOI: 10.1016/j.molcel.2008.12.029
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ABSTRACT The mixed-lineage leukemia protein MLL1 is a transcriptional regulator with an essential role in early development and hematopoiesis. The biological function of MLL1 is mediated by the histone H3K4 methyltransferase activity of the carboxyl-terminal SET domain. We have determined the crystal structure of the MLL1 SET domain in complex with cofactor product AdoHcy and a histone H3 peptide. This structure indicates that, in order to form a well-ordered active site, a highly variable but essential component of the SET domain must be repositioned. To test this idea, we compared the effect of the addition of MLL complex members on methyltransferase activity and show that both RbBP5 and Ash2L but not Wdr5 stimulate activity. Additionally, we have determined the effect of posttranslational modifications on histone H3 residues downstream and upstream from the target lysine and provide a structural explanation for why H3T3 phosphorylation and H3K9 acetylation regulate activity.

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    • "All ternary structures of SET domain methyltransferases in complex with cofactor and substrate have the substrate peptide lying in a narrow groove formed by the I-SET domain on one side, and the post-SET domain on the opposite side. This peptide binding groove is responsible for substrate sequence specificity and positions the substrate lysine side chain into a conserved channel that goes deep into the core of the SET domain and meets the cofactor at the conserved active site as described above (Figure 2) ([38,43,44,47,48,49,50,51,52] and PDB code 4AU7). The width of the peptide-binding groove is critical for catalysis, and the distance separating the two edges is between 7.2 and 7.8 Å across all available structures of active SET domain constructs (see materials and methods section for details). "
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    ABSTRACT: Polycomb repressive complex 2 (PRC2) is an important regulator of cellular differentiation and cell type identity. Overexpression or activating mutations of EZH2, the catalytic component of the PRC2 complex, are linked to hyper-trimethylation of lysine 27 of histone H3 (H3K27me3) in many cancers. Potent EZH2 inhibitors that reduce levels of H3K27me3 kill mutant lymphoma cells and are efficacious in a mouse xenograft model of malignant rhabdoid tumors. Unlike most SET domain methyltransferases, EZH2 requires PRC2 components, SUZ12 and EED, for activity, but the mechanism by which catalysis is promoted in the PRC2 complex is unknown. We solved the 2.0 Å crystal structure of the EZH2 methyltransferase domain revealing that most of the canonical structural features of SET domain methyltransferase structures are conserved. The site of methyl transfer is in a catalytically competent state, and the structure clarifies the structural mechanism underlying oncogenic hyper-trimethylation of H3K27 in tumors harboring mutations at Y641 or A677. On the other hand, the I-SET and post-SET domains occupy atypical positions relative to the core SET domain resulting in incomplete formation of the cofactor binding site and occlusion of the substrate binding groove. A novel CXC domain N-terminal to the SET domain may contribute to the apparent inactive conformation. We propose that protein interactions within the PRC2 complex modulate the trajectory of the post-SET and I-SET domains of EZH2 in favor of a catalytically competent conformation.
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    • "Structure of the Autoinhibited PRDM9 PR/SET Domain The post-SET domain is disordered in the unbound mPRDM9 molecule in the crystal (Figure S1B). Similarly, the post-SET domains of DIM-5 or MLL1 become fully folded only in the presence of the AdoHcy/AdoMet cofactor and substrate (Southall et al., 2009; Zhang et al., 2002). To better understand the role of the post-SET domain, we produced crystals of a larger fragment of mouse PRDM9 that, however, only diffracted to a low resolution. "
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    ABSTRACT: PRDM9, a histone lysine methyltransferase, is a key determinant of the localization of meiotic recombination hot spots in humans and mice and the only vertebrate protein known to be involved in hybrid sterility. Here, we report the crystal structure of the PRDM9 methyltransferase domain in complex with a histone H3 peptide dimethylated on lysine 4 (H3K4me2) and S-adenosylhomocysteine (AdoHcy), which provides insights into the methyltransferase activity of PRDM proteins. We show that the genuine substrate of PRDM9 is histone H3 lysine 4 (H3K4) and that the enzyme possesses mono-, di-, and trimethylation activities. We also determined the crystal structure of PRDM9 in its autoinhibited state, which revealed a rearrangement of the substrate and cofactor binding sites by a concerted action of the pre-SET and post-SET domains, providing important insights into the regulatory mechanisms of histone lysine methyltransferase activity.
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    • "MLL is homologous to the Drosophila trithorax gene and both protein products play important roles in epigenetics by perpetuating the chromatin structure through propagating pre‐ set gene expression signatures, thus conveying the epigenetic information to the progeny. MLL is part of large chromatin modifier complexes and entails histone methyltransferase activity that modifies histone H3 lysine 4 (H3K4) and H3 lysine 79 (H3K79) that leads to gene activation and transcription elongation, such as of HOX genes that are crucial for organization of the body axis and cell lineage definition (Southall et al, 2009). Early demonstration of B‐cell to myeloid reprogramming was achieved by retroviral encoded signalling oncogenes, such as Ras or Raf that also activate C/EBPb (Klinken et al, 1988). "
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