A Conserved Arginine-containing Motif Crucial for the Assembly and Enzymatic Activity of the Mixed Lineage Leukemia Protein-1 Core Complex
ABSTRACT The mixed lineage leukemia protein-1 (MLL1) belongs to the SET1 family of histone H3 lysine 4 methyltransferases. Recent studies indicate that the catalytic subunits of SET1 family members are regulated by interaction with a conserved core group of proteins that include the WD repeat protein-5 (WDR5), retinoblastoma-binding protein-5 (RbBP5), and the absent small homeotic-2-like protein (Ash2L). It has been suggested that WDR5 functions to bridge the interactions between the catalytic and regulatory subunits of SET1 family complexes. However, the molecular details of these interactions are unknown. To gain insight into the interactions among these proteins, we have determined the biophysical basis for the interaction between the human WDR5 and MLL1. Our studies reveal that WDR5 preferentially recognizes a previously unidentified and conserved arginine-containing motif, called the "Win" or WDR5 interaction motif, which is located in the N-SET region of MLL1 and other SET1 family members. Surprisingly, our structural and functional studies show that WDR5 recognizes arginine 3765 of the MLL1 Win motif using the same arginine binding pocket on WDR5 that was previously shown to bind histone H3. We demonstrate that WDR5's recognition of arginine 3765 of MLL1 is essential for the assembly and enzymatic activity of the MLL1 core complex in vitro.
- SourceAvailable from: Venkat Dharmarajan
Acute Leukemia - The Scientist's Perspective and Challenge, 12/2011; , ISBN: 978-953-307-553-2
- "The extensive biochemical characterization of the peptide derived from the MLL1 Win motif have demonstrated that these peptidomimetic compounds have the potential to inhibit the H3K4 methyltransferase activity of the MLL1 core complex by specifically disrupting the association of MLL1 with the WRAD sub-complex (Patel et al., 2008b). Furthermore, the crystal structures of WDR5 bound to human SET1 family Win motif peptides provides a rationale for the design of new peptides and non-peptide mimetics with better inhibitory properties. "
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- "These results demonstrate that CHD8 has extensive contacts with the WAR subcomplex, unlike the interactions with MLL that are dependent on WDR5    . The precise means by which these interactions occur, including interacting domains between each protein, remains to be elucidated. "
ABSTRACT: Chromodomain, helicase, DNA-binding protein 8 (CHD8) is an ATP-dependent chromatin remodeling enzyme that has been demonstrated to exist within a large protein complex which includes WDR5, Ash2L, and RbBP5, members of the Mixed Lineage Leukemia (MLL) histone modifying complexes. Here we show that CHD8 relocalizes to the promoter of the MLL regulated gene HOXA2 upon gene activation. Depletion of CHD8 enhances HOXA2 expression under activating conditions. Furthermore, depletion of CHD8 results in a loss of the WDR5/Ash2L/RbBP5 subcomplex, and consequently H3K4 trimethylation, at the HOXA2 promoter. These studies suggest that CHD8 alters HOXA2 gene expression and regulates the recruitment of chromatin modifying enzymes.FEBS letters 02/2010; 584(4):689-93. DOI:10.1016/j.febslet.2010.01.022 · 3.34 Impact Factor
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ABSTRACT: ATP-dependent chromatin remodeling by the CHD family of proteins plays an important role in the regulation of gene transcription. The CHD family can be subdivided into three families; CHD1-2, CHD3-5, and CHD6-9. While the first two subfamilies have been extensively studied, very little is known about the CHD6-9 subfamily. In this study we demonstrate that CHD8 is a nucleosome-stimulated ATPase, capable of remodeling nucleosome structure. In addition, the tandem Chromodomains of CHD8 are capable of directly binding recombinant histones H3 and H4. We also demonstrate that CHD8 interacts directly with the transcriptional regulator β-catenin and that CHD8 is recruited specifically to the promoter regions of several genes responsive to β-catenin. Utilizing shRNA against CHD8, we demonstrate that CHD8 performs a negative role in regulating β-catenin target gene expression. This regulation is evolutionarily conserved as RNAi against kismet, the apparent Drosophila ortholog of CHD8, similarly results in the activation of β-catenin target genes. WDR5, RbBP5, and Ash2L are core components of the MLL1 WDR5 methyltransferase complex which alters chromatin structure through the covalent modification of histones. MLL1, the catalytic subunit of the complex, catalyzes the methylation of histone H3 lysine 4, a hallmark of active chromatin. We demonstrate that CHD8 exists in a multisubunit complex with WDR5, RbBP5, and Ash2L that may also contain MLL1. Both WDR5 and MLL1 have previously been reported to regulate the expression of Hox genes, a family of genes involved in development. We demonstrate that CHD8 is recruited specifically to the promoter regions of several genes within the HoxA locus. Utilizing shRNA against CHD8, we demonstrate that CHD8 performs a negative role in regulating Hox gene expression. We show that CHD8, like WDR5 and MLL1, regulates Hox gene expression. Taken together, these results demonstrate that CHD8 functions in the transcriptional regulation of both β-catenin target genes and Hox genes and suggest that this regulation is through the ATP-dependent modulation of chromatin structure within the 5’ promoter regions of these genes. Our results suggest that through regulating the expression of β-catenin target genes and Hox genes CHD8 may play a role in both tumorogenesis and development respectively. Ph.D. Cellular & Molecular Biology University of Michigan, Horace H. Rackham School of Graduate Studies http://deepblue.lib.umich.edu/bitstream/2027.42/62235/1/bathomps_1.pdf