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The plasticity of WDR5 peptide-binding cleft enables the binding of the SET1 family of histone methyltransferases.

Zhang P, Lee H, Brunzelle JS, Couture JF - Nucleic Acids Res. (2012)

Bottom Line: We herein provide biochemical and structural evidence showing that WDR5 binds the Win motifs of MLL2-4, SET1A and SET1B.Comparative analysis of WDR5-Win complexes reveals that binding of the Win motifs is achieved by the plasticity of WDR5 peptidyl-arginine-binding cleft allowing the C-terminal ends of the Win motifs to be maintained in structurally divergent conformations.Overall, our findings illustrate the function of WDR5 in scaffolding the SET1 family of KMTs and further emphasize on the important role of WDR5 in regulating global histone H3 Lys-4 methylation.

View Article: PubMed Central - PubMed

Affiliation: Ottawa Institute of Systems Biology, Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada.

ABSTRACT
In mammals, the SET1 family of lysine methyltransferases (KMTs), which includes MLL1-5, SET1A and SET1B, catalyzes the methylation of lysine-4 (Lys-4) on histone H3. Recent reports have demonstrated that a three-subunit complex composed of WD-repeat protein-5 (WDR5), retinoblastoma-binding protein-5 (RbBP5) and absent, small, homeotic disks-2-like (ASH2L) stimulates the methyltransferase activity of MLL1. On the basis of studies showing that this stimulation is in part controlled by an interaction between WDR5 and a small region located in close proximity of the MLL1 catalytic domain [referred to as the WDR5-interacting motif (Win)], it has been suggested that WDR5 might play an analogous role in scaffolding the other SET1 complexes. We herein provide biochemical and structural evidence showing that WDR5 binds the Win motifs of MLL2-4, SET1A and SET1B. Comparative analysis of WDR5-Win complexes reveals that binding of the Win motifs is achieved by the plasticity of WDR5 peptidyl-arginine-binding cleft allowing the C-terminal ends of the Win motifs to be maintained in structurally divergent conformations. Consistently, enzymatic assays reveal that WDR5 plays an important role in the optimal stimulation of MLL2-4, SET1A and SET1B methyltransferase activity by the RbBP5-ASH2L heterodimer. Overall, our findings illustrate the function of WDR5 in scaffolding the SET1 family of KMTs and further emphasize on the important role of WDR5 in regulating global histone H3 Lys-4 methylation.

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WDR5 binds MLL1-4, SET1A and SET1B Win motifs. ITC titration experiments with MLL1Win (A), MLL2Win (B), MLL3Win (C), MLL4Win (D), SET1AWin (E) and SET1BWin (F) peptides and WDR5 (upper trace) and the fitted binding curve (lower trace). The sequence of the Win motifs and the KD are indicated as insets. Titration of WDR5 with Win peptides displays binding stoichiometries (N values) between 0.9 and 1.0.
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gkr1235-F1: WDR5 binds MLL1-4, SET1A and SET1B Win motifs. ITC titration experiments with MLL1Win (A), MLL2Win (B), MLL3Win (C), MLL4Win (D), SET1AWin (E) and SET1BWin (F) peptides and WDR5 (upper trace) and the fitted binding curve (lower trace). The sequence of the Win motifs and the KD are indicated as insets. Titration of WDR5 with Win peptides displays binding stoichiometries (N values) between 0.9 and 1.0.

Mentions: A recent biochemical analysis of WDR5 peptidyl-arginine-binding cleft revealed that this protein binds a 10-residue peptide corresponding to MLL1Win with an equilibrium dissociation constant (KD) of 1.7 µM. The crystal structure of WDR5 in complex with MLL1Win identified that R3765 of MLL1 was critical in conferring binding to WDR5, scaffolding the MLL1–WDR5–RbBP5–ASH2L complex and, correlatively, stimulation of MLL1 methyltransferase activity (23). Accordingly, the same authors posited that a similar region found on other members of the SET1 family would play an analogous role (31) while another study proposed that only MLL1 and MLL4 could bind WDR5 (30). In order to investigate these hypotheses, we first sought to measure the KD values of WDR5 for MLL1-4 and SET1A/BWin motifs. Using ITC, we determined that WDR5 binds MLL1 Win and MLL4Win peptides with a KD of 1.13 µM and 0.03 µM, respectively. Similar titration experiments of WDR5 with MLL2Win and MLL3Win peptides resulted in KD of 0.16 µM and 2.03 µM, respectively, while SET1AWin and SET1BWin peptides bound to WDR5 with a KD of 0.26 µM and 0.10 µM, respectively. Overall, these results demonstrate that WDR5 binds the Win region of all the members of the SET1 family of KMTs. However, given the divergence in the amino acid sequence succeeding the conserved arginine residue and the ∼37-fold range of binding affinities of WDR5 for the Win motifs (Table 1 and Figure 1), our binding analysis suggests that WDR5 interacts with SET1 KMTs using divergent modes.Figure 1.


The plasticity of WDR5 peptide-binding cleft enables the binding of the SET1 family of histone methyltransferases.

Zhang P, Lee H, Brunzelle JS, Couture JF - Nucleic Acids Res. (2012)

WDR5 binds MLL1-4, SET1A and SET1B Win motifs. ITC titration experiments with MLL1Win (A), MLL2Win (B), MLL3Win (C), MLL4Win (D), SET1AWin (E) and SET1BWin (F) peptides and WDR5 (upper trace) and the fitted binding curve (lower trace). The sequence of the Win motifs and the KD are indicated as insets. Titration of WDR5 with Win peptides displays binding stoichiometries (N values) between 0.9 and 1.0.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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gkr1235-F1: WDR5 binds MLL1-4, SET1A and SET1B Win motifs. ITC titration experiments with MLL1Win (A), MLL2Win (B), MLL3Win (C), MLL4Win (D), SET1AWin (E) and SET1BWin (F) peptides and WDR5 (upper trace) and the fitted binding curve (lower trace). The sequence of the Win motifs and the KD are indicated as insets. Titration of WDR5 with Win peptides displays binding stoichiometries (N values) between 0.9 and 1.0.
Mentions: A recent biochemical analysis of WDR5 peptidyl-arginine-binding cleft revealed that this protein binds a 10-residue peptide corresponding to MLL1Win with an equilibrium dissociation constant (KD) of 1.7 µM. The crystal structure of WDR5 in complex with MLL1Win identified that R3765 of MLL1 was critical in conferring binding to WDR5, scaffolding the MLL1–WDR5–RbBP5–ASH2L complex and, correlatively, stimulation of MLL1 methyltransferase activity (23). Accordingly, the same authors posited that a similar region found on other members of the SET1 family would play an analogous role (31) while another study proposed that only MLL1 and MLL4 could bind WDR5 (30). In order to investigate these hypotheses, we first sought to measure the KD values of WDR5 for MLL1-4 and SET1A/BWin motifs. Using ITC, we determined that WDR5 binds MLL1 Win and MLL4Win peptides with a KD of 1.13 µM and 0.03 µM, respectively. Similar titration experiments of WDR5 with MLL2Win and MLL3Win peptides resulted in KD of 0.16 µM and 2.03 µM, respectively, while SET1AWin and SET1BWin peptides bound to WDR5 with a KD of 0.26 µM and 0.10 µM, respectively. Overall, these results demonstrate that WDR5 binds the Win region of all the members of the SET1 family of KMTs. However, given the divergence in the amino acid sequence succeeding the conserved arginine residue and the ∼37-fold range of binding affinities of WDR5 for the Win motifs (Table 1 and Figure 1), our binding analysis suggests that WDR5 interacts with SET1 KMTs using divergent modes.Figure 1.

Bottom Line: We herein provide biochemical and structural evidence showing that WDR5 binds the Win motifs of MLL2-4, SET1A and SET1B.Comparative analysis of WDR5-Win complexes reveals that binding of the Win motifs is achieved by the plasticity of WDR5 peptidyl-arginine-binding cleft allowing the C-terminal ends of the Win motifs to be maintained in structurally divergent conformations.Overall, our findings illustrate the function of WDR5 in scaffolding the SET1 family of KMTs and further emphasize on the important role of WDR5 in regulating global histone H3 Lys-4 methylation.

View Article: PubMed Central - PubMed

Affiliation: Ottawa Institute of Systems Biology, Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada.

ABSTRACT
In mammals, the SET1 family of lysine methyltransferases (KMTs), which includes MLL1-5, SET1A and SET1B, catalyzes the methylation of lysine-4 (Lys-4) on histone H3. Recent reports have demonstrated that a three-subunit complex composed of WD-repeat protein-5 (WDR5), retinoblastoma-binding protein-5 (RbBP5) and absent, small, homeotic disks-2-like (ASH2L) stimulates the methyltransferase activity of MLL1. On the basis of studies showing that this stimulation is in part controlled by an interaction between WDR5 and a small region located in close proximity of the MLL1 catalytic domain [referred to as the WDR5-interacting motif (Win)], it has been suggested that WDR5 might play an analogous role in scaffolding the other SET1 complexes. We herein provide biochemical and structural evidence showing that WDR5 binds the Win motifs of MLL2-4, SET1A and SET1B. Comparative analysis of WDR5-Win complexes reveals that binding of the Win motifs is achieved by the plasticity of WDR5 peptidyl-arginine-binding cleft allowing the C-terminal ends of the Win motifs to be maintained in structurally divergent conformations. Consistently, enzymatic assays reveal that WDR5 plays an important role in the optimal stimulation of MLL2-4, SET1A and SET1B methyltransferase activity by the RbBP5-ASH2L heterodimer. Overall, our findings illustrate the function of WDR5 in scaffolding the SET1 family of KMTs and further emphasize on the important role of WDR5 in regulating global histone H3 Lys-4 methylation.

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