Structural characterization of Set1 RNA recognition motifs and their role in histone H3 lysine 4 methylation.

The yeast Set1 histone H3 lysine 4 (H3K4) methyltransferase contains, in addition to its catalytic SET domain, a conserved RNA recognition motif (RRM1). We present here the crystal structure and the secondary structure assignment in solution of the Set1 RRM1. Although RRM1 has the expected betaalphabetabetaalphabeta RRM-fold, it lacks the typical RNA-binding features of these modules. RRM1 is not able to bind RNA by itself in vitro, but a construct combining RRM1 with a newly identified downstream RRM2 specifically binds RNA. In vivo, H3K4 methylation is not affected by a point mutation in RRM2 that preserves Set1 stability but affects RNA binding in vitro. In contrast mutating RRM1 destabilizes Set1 and leads to an increase of dimethylation of H3K4 at the 5'-coding region of active genes at the expense of trimethylation, whereas both, dimethylation decreases at the 3'-coding region. Taken together, our results suggest that Set1 RRMs bind RNA, but Set1 RNA-binding activity is not linked to H3K4 methylation.

[1]  Kevin Struhl,et al.  Targeted recruitment of Set1 histone methylase by elongating Pol II provides a localized mark and memory of recent transcriptional activity. , 2003, Molecular cell.

[2]  Arne Elofsson,et al.  3D-Jury: A Simple Approach to Improve Protein Structure Predictions , 2003, Bioinform..

[3]  Marcin von Grotthuss,et al.  Detecting distant homology with Meta-BASIC , 2004, Nucleic Acids Res..

[4]  R. Kornberg,et al.  A trithorax-group complex purified from Saccharomyces cerevisiae is required for methylation of histone H3 , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[5]  Mark Johnston,et al.  The Paf1 Complex Is Essential for Histone Monoubiquitination by the Rad6-Bre1 Complex, Which Signals for Histone Methylation by COMPASS and Dot1p* , 2003, Journal of Biological Chemistry.

[6]  T. Hughes,et al.  BUR Kinase Selectively Regulates H3 K4 Trimethylation and H2B Ubiquitylation through Recruitment of the PAF Elongation Complex , 2005, Current Biology.

[7]  Mark Johnston,et al.  Methylation of Histone H3 by COMPASS Requires Ubiquitination of Histone H2B by Rad6* , 2002, The Journal of Biological Chemistry.

[8]  Nevan J. Krogan,et al.  COMPASS: A complex of proteins associated with a trithorax-related SET domain protein , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[9]  Rein Aasland,et al.  The Saccharomyces cerevisiae Set1 complex includes an Ash2 homologue and methylates histone 3 lysine 4 , 2001, The EMBO journal.

[10]  Patrice Gouet,et al.  ESPript: analysis of multiple sequence alignments in PostScript , 1999, Bioinform..

[11]  S Cusack,et al.  Crystal structure of the human nuclear cap binding complex. , 2001, Molecular cell.

[12]  Ali Shilatifard,et al.  Transcriptional activation via sequential histone H2B ubiquitylation and deubiquitylation, mediated by SAGA-associated Ubp8. , 2003, Genes & development.

[13]  Megan F. Cole,et al.  Genome-wide Map of Nucleosome Acetylation and Methylation in Yeast , 2005, Cell.

[14]  T. Jenuwein,et al.  The many faces of histone lysine methylation. , 2002, Current opinion in cell biology.

[15]  Thomas L. Madden,et al.  Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. , 1997, Nucleic acids research.

[16]  Ian M. Fingerman,et al.  Global Loss of Set1-mediated H3 Lys4 Trimethylation Is Associated with Silencing Defects in Saccharomyces cerevisiae* , 2005, Journal of Biological Chemistry.

[17]  Bradley R Cairns,et al.  Histone trimethylation by Set1 is coordinated by the RRM, autoinhibitory, and catalytic domains , 2005, The EMBO journal.

[18]  Nathan A. Baker,et al.  Electrostatics of nanosystems: Application to microtubules and the ribosome , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[19]  Zu-Wen Sun,et al.  Ubiquitination of histone H2B regulates H3 methylation and gene silencing in yeast , 2002, Nature.

[20]  G. Murshudov,et al.  Refinement of macromolecular structures by the maximum-likelihood method. , 1997, Acta crystallographica. Section D, Biological crystallography.

[21]  Michael Hampsey,et al.  Tails of Intrigue Phosphorylation of RNA Polymerase II Mediates Histone Methylation , 2003, Cell.

[22]  F. Ishikawa,et al.  Structure and interactions with RNA of the N-terminal UUAG-specific RNA-binding domain of hnRNP D0. , 1999, Journal of molecular biology.

[23]  A. Shilatifard,et al.  Molecular regulation of histone H3 trimethylation by COMPASS and the regulation of gene expression. , 2005, Molecular cell.

[24]  P. Evans,et al.  Crystal structure of the RNA-binding domain of the U1 small nuclear ribonucleoprotein A , 1990, Nature.

[25]  Rodrigo Lopez,et al.  Multiple sequence alignment with the Clustal series of programs , 2003, Nucleic Acids Res..

[26]  Liam J. McGuffin,et al.  Improvement of the GenTHREADER Method for Genomic Fold Recognition , 2003, Bioinform..

[27]  J. Davie,et al.  Histone H3 lysine 4 methylation is mediated by Set1 and required for cell growth and rDNA silencing in Saccharomyces cerevisiae. , 2001, Genes & development.

[28]  G. Bricogne,et al.  [27] Maximum-likelihood heavy-atom parameter refinement for multiple isomorphous replacement and multiwavelength anomalous diffraction methods. , 1997, Methods in enzymology.

[29]  Kevin Struhl,et al.  The Rtf1 Component of the Paf1 Transcriptional Elongation Complex Is Required for Ubiquitination of Histone H2B* , 2003, Journal of Biological Chemistry.

[30]  M. Johnston,et al.  The Paf1 complex is required for histone H3 methylation by COMPASS and Dot1p: linking transcriptional elongation to histone methylation. , 2003, Molecular cell.

[31]  T. Blundell,et al.  Comparative protein modelling by satisfaction of spatial restraints. , 1993, Journal of molecular biology.

[32]  T L Blundell,et al.  FUGUE: sequence-structure homology recognition using environment-specific substitution tables and structure-dependent gap penalties. , 2001, Journal of molecular biology.

[33]  J. Mellor,et al.  Dynamic lysine methylation on histone H3 defines the regulatory phase of gene transcription. , 2005, Molecular cell.

[34]  D. van der Spoel,et al.  GROMACS: A message-passing parallel molecular dynamics implementation , 1995 .

[35]  Thomas C Terwilliger,et al.  Automated structure solution, density modification and model building. , 2002, Acta crystallographica. Section D, Biological crystallography.

[36]  S. Cusack,et al.  Co-crystallization of the human nuclear cap-binding complex with a m7GpppG cap analogue using protein engineering. , 2002, Acta crystallographica. Section D, Biological crystallography.

[37]  P. Evans,et al.  The RNP domain: a sequence-specific RNA-binding domain involved in processing and transport of RNA. , 1995, Trends in biochemical sciences.

[38]  G. Labesse,et al.  Deciphering protein sequence information through hydrophobic cluster analysis (HCA): current status and perspectives , 1997, Cellular and Molecular Life Sciences CMLS.

[39]  Thomas C. Terwilliger,et al.  Automated MAD and MIR structure solution , 1999, Acta crystallographica. Section D, Biological crystallography.

[40]  Mark Johnston,et al.  The Bur1/Bur2 complex is required for histone H2B monoubiquitination by Rad6/Bre1 and histone methylation by COMPASS. , 2005, Molecular cell.

[41]  Stuart L. Schreiber,et al.  Active genes are tri-methylated at K4 of histone H3 , 2002, Nature.

[42]  Alain Verreault,et al.  Histone H3 lysine 4 mono-methylation does not require ubiquitination of histone H2B. , 2005, Journal of molecular biology.

[43]  D. Bentley,et al.  Altered nucleosome occupancy and histone H3K4 methylation in response to ‘transcriptional stress’ , 2005, The EMBO journal.

[44]  E. Ezhkova,et al.  Proteasomal ATPases link ubiquitylation of histone H2B to methylation of histone H3. , 2004, Molecular cell.

[45]  Michael Grunstein,et al.  Histone H2B ubiquitylation controls processive methylation but not monomethylation by Dot1 and Set1. , 2005, Molecular cell.

[46]  Collaborative Computational,et al.  The CCP4 suite: programs for protein crystallography. , 1994, Acta crystallographica. Section D, Biological crystallography.

[47]  K Henrick,et al.  Electronic Reprint Biological Crystallography Secondary-structure Matching (ssm), a New Tool for Fast Protein Structure Alignment in Three Dimensions Biological Crystallography Secondary-structure Matching (ssm), a New Tool for Fast Protein Structure Alignment in Three Dimensions , 2022 .

[48]  George M Sheldrick,et al.  Substructure solution with SHELXD. , 2002, Acta crystallographica. Section D, Biological crystallography.

[49]  K. Struhl,et al.  Binding of TBP to promoters in vivo is stimulated by activators and requires Pol II holoenzyme , 1999, Nature.