The Human Mixed Lineage Leukemia 5 (MLL5), a Sequentially and Structurally Divergent SET Domain-Containing Protein with No Intrinsic Catalytic Activity

Mixed Lineage Leukemia 5 (MLL5) plays a key role in hematopoiesis, spermatogenesis and cell cycle progression. Chromatin binding is ensured by its plant homeodomain (PHD) through a direct interaction with the N-terminus of histone H3 (H3). In addition, MLL5 contains a Su(var)3-9, Enhancer of zeste, Trithorax (SET) domain, a protein module that usually displays histone lysine methyltransferase activity. We report here the crystal structure of the unliganded SET domain of human MLL5 at 2.1 Å resolution. Although it shows most of the canonical features of other SET domains, both the lack of key residues and the presence in the SET-I subdomain of an unusually large loop preclude the interaction of MLL5 SET with its cofactor and substrate. Accordingly, we show that MLL5 is devoid of any in vitro methyltransferase activity on full-length histones and histone H3 peptides. Hence, the three dimensional structure of MLL5 SET domain unveils the structural basis for its lack of methyltransferase activity and suggests a new regulatory mechanism.

[1]  Ben M. Webb,et al.  Comparative Protein Structure Modeling Using MODELLER , 2016, Current protocols in bioinformatics.

[2]  Jon R. Wilson,et al.  Structural basis of oncogenic histone H3K27M inhibition of human polycomb repressive complex 2 , 2016, Nature Communications.

[3]  Sudhir Kumar,et al.  MEGA7: Molecular Evolutionary Genetics Analysis Version 7.0 for Bigger Datasets. , 2016, Molecular biology and evolution.

[4]  Shuai Li,et al.  Structural basis for activity regulation of MLL family methyltransferases , 2016, Nature.

[5]  Lars Skjærven,et al.  WEBnm@ v2.0: Web server and services for comparing protein flexibility , 2014, BMC Bioinformatics.

[6]  C. Arrowsmith,et al.  Structure of the Catalytic Domain of EZH2 Reveals Conformational Plasticity in Cofactor and Substrate Binding Sites and Explains Oncogenic Mutations , 2013, PloS one.

[7]  J. Bonanno,et al.  Structural Context of Disease-Associated Mutations and Putative Mechanism of Autoinhibition Revealed by X-Ray Crystallographic Analysis of the EZH2-SET Domain , 2013, PloS one.

[8]  S. Aparicio,et al.  Solution NMR Structure and Histone Binding of the PHD Domain of Human MLL5 , 2013, PloS one.

[9]  Baldomero Oliva,et al.  MODELLER: A Program for Protein Structure Modeling , 2013 .

[10]  M. Groudine,et al.  Molecular basis for chromatin binding and regulation of MLL5 , 2013, Proceedings of the National Academy of Sciences.

[11]  Chuangui Wang,et al.  Mixed Lineage Leukemia 5 (MLL5) Protein Regulates Cell Cycle Progression and E2F1-responsive Gene Expression via Association with Host Cell Factor-1 (HCF-1)* , 2013, The Journal of Biological Chemistry.

[12]  A. Shilatifard,et al.  Histone H 3 Lysine 4 ( H 3 K 4 ) Methylation in Development and Differentiation , 2013 .

[13]  M. Groudine,et al.  UpSET Recruits HDAC Complexes and Restricts Chromatin Accessibility and Acetylation at Promoter Regions , 2012, Cell.

[14]  J. Min,et al.  Sinefungin derivatives as inhibitors and structure probes of protein lysine methyltransferase SETD2. , 2012, Journal of the American Chemical Society.

[15]  P. Zwart,et al.  Towards automated crystallographic structure refinement with phenix.refine , 2012, Acta crystallographica. Section D, Biological crystallography.

[16]  Y. G. Zheng,et al.  Scintillation Proximity Assay of Arginine Methylation , 2012, Journal of biomolecular screening.

[17]  J. Couture,et al.  The plasticity of WDR5 peptide-binding cleft enables the binding of the SET1 family of histone methyltransferases , 2012, Nucleic acids research.

[18]  G. Turashvili,et al.  Mll5 Is Required for Normal Spermatogenesis , 2011, PloS one.

[19]  H. Al‐Hashimi,et al.  Direct Evidence for Methyl Group Coordination by Carbon-Oxygen Hydrogen Bonds in the Lysine Methyltransferase SET7/9* , 2011, The Journal of Biological Chemistry.

[20]  Randy J. Read,et al.  Overview of the CCP4 suite and current developments , 2011, Acta crystallographica. Section D, Biological crystallography.

[21]  Liisa Holm,et al.  Dali server: conservation mapping in 3D , 2010, Nucleic Acids Res..

[22]  P. Emsley,et al.  Features and development of Coot , 2010, Acta crystallographica. Section D, Biological crystallography.

[23]  Kevin Cowtan,et al.  Recent developments in classical density modification , 2010, Acta crystallographica. Section D, Biological crystallography.

[24]  A. Shilatifard,et al.  Histone H3 lysine 4 (H3K4) methylation in development and differentiation. , 2010, Developmental biology.

[25]  Randy J. Read,et al.  Acta Crystallographica Section D Biological , 2003 .

[26]  S. Buratowski,et al.  Dimethylation of H3K4 by Set1 Recruits the Set3 Histone Deacetylase Complex to 5′ Transcribed Regions , 2009, Cell.

[27]  G. Pavlath,et al.  MLL5, a trithorax homolog, indirectly regulates H3K4 methylation, represses cyclin A2 expression, and promotes myogenic differentiation , 2009, Proceedings of the National Academy of Sciences.

[28]  S. Aparicio,et al.  Loss of MLL5 results in pleiotropic hematopoietic defects, reduced neutrophil immune function, and extreme sensitivity to DNA demethylation. , 2009, Blood.

[29]  N. Killeen,et al.  MLL5 contributes to hematopoietic stem cell fitness and homeostasis. , 2009, Blood.

[30]  K. Döhner,et al.  Impaired function of primitive hematopoietic cells in mice lacking the Mixed-Lineage-Leukemia homolog MLL5. , 2009, Blood.

[31]  Jon R. Wilson,et al.  Structural basis for the requirement of additional factors for MLL1 SET domain activity and recognition of epigenetic marks. , 2009, Molecular cell.

[32]  M. Cosgrove,et al.  Structure of WDR5 Bound to Mixed Lineage Leukemia Protein-1 Peptide* , 2008, Journal of Biological Chemistry.

[33]  O. Gascuel,et al.  An improved general amino acid replacement matrix. , 2008, Molecular biology and evolution.

[34]  A. Shilatifard.  Molecular implementation and physiological roles for histone H3 lysine 4 (H3K4) methylation. , 2008, Current opinion in cell biology.

[35]  Randy J. Read,et al.  Phaser crystallographic software , 2007, Journal of applied crystallography.

[36]  David Alderton,et al.  A versatile ligation-independent cloning method suitable for high-throughput expression screening applications , 2007, Nucleic acids research.

[37]  C. Allis,et al.  Methylation of lysine 4 on histone H3: intricacy of writing and reading a single epigenetic mark. , 2007, Molecular cell.

[38]  Kevin Cowtan,et al.  The Buccaneer software for automated model building. 1. Tracing protein chains. , 2006, Acta crystallographica. Section D, Biological crystallography.

[39]  Laxmikant V. Kalé,et al.  Scalable molecular dynamics with NAMD , 2005, J. Comput. Chem..

[40]  Fei Long,et al.  REFMAC5 dictionary: organization of prior chemical knowledge and guidelines for its use. , 2004, Acta crystallographica. Section D, Biological crystallography.

[41]  Stephen H. Bryant,et al.  CD-Search: protein domain annotations on the fly , 2004, Nucleic Acids Res..

[42]  J. Strominger,et al.  MLL 5 protein forms intranuclear foci, and overexpression inhibits cell cycle progression. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[43]  Geoff Kelly,et al.  Structure and catalytic mechanism of the human histone methyltransferase SET7/9 , 2003, Nature.

[44]  E. Green,et al.  MLL5, a homolog of Drosophila trithorax located within a segment of chromosome band 7q22 implicated in myeloid leukemia , 2002, Oncogene.

[45]  M Wilm,et al.  The S. cerevisiae SET3 complex includes two histone deacetylases, Hos2 and Hst1, and is a meiotic-specific repressor of the sporulation gene program. , 2001, Genes & development.

[46]  L. Nilsson,et al.  Structure and Dynamics of the TIP3P, SPC, and SPC/E Water Models at 298 K , 2001 .

[47]  John P. Huelsenbeck,et al.  MRBAYES: Bayesian inference of phylogenetic trees , 2001, Bioinform..

[48]  P. Lio’,et al.  Molecular phylogenetics: state-of-the-art methods for looking into the past. , 2001, Trends in genetics : TIG.

[49]  P. Hünenberger,et al.  A fast SHAKE algorithm to solve distance constraint equations for small molecules in molecular dynamics simulations , 2001, J. Comput. Chem..

[50]  P Mark,et al.  298KでのTIP3P,SPC及びSPC/E水モデルの構造及び動力学 , 2001 .

[51]  J. Huelsenbeck,et al.  MRBAYES : Bayesian inference of phylogeny , 2001 .

[52]  C D Kroenke,et al.  Nuclear magnetic resonance methods for quantifying microsecond-to-millisecond motions in biological macromolecules. , 2001, Methods in enzymology.

[53]  Bernd Meyer,et al.  Characterization of Ligand Binding by Saturation Transfer Difference NMR Spectroscopy. , 1999, Angewandte Chemie.

[54]  Alexander D. MacKerell,et al.  All-atom empirical potential for molecular modeling and dynamics studies of proteins. , 1998, The journal of physical chemistry. B.

[55]  T. Malliavin,et al.  Gifa V. 4: A complete package for NMR data set processing , 1996, Journal of biomolecular NMR.

[56]  K Schulten,et al.  VMD: visual molecular dynamics. , 1996, Journal of molecular graphics.

[57]  T. Darden,et al.  Particle mesh Ewald: An N⋅log(N) method for Ewald sums in large systems , 1993 .

[58]  J. Felsenstein.  CONFIDENCE LIMITS ON PHYLOGENIES: AN APPROACH USING THE BOOTSTRAP , 1985, Evolution; international journal of organic evolution.

[59]  Ballard,et al.  Overview of the CCP 4 suite and current developments , 2022 .