Rui Ming Xu, Ph.D.

The structure of NSD1 reveals an autoregulatory mechanism underlying histone H3K36 methylation
Qiao, Qi; Li, Yan; Chen, Zhi; Wang, Mingzhu; Reinberg, Danny; Xu, Rui-Ming
2011 Mar 11;286(10):8361-8368, Journal of biological chemistry
The Sotos syndrome gene product, NSD1, is a SET domain histone methyltransferase that primarily dimethylates nucleosomal histone H3 lysine 36 (H3K36). To date, the intrinsic properties of NSD1 that determine its nucleosomal substrate selectivity and dimethyl H3K36 product specificity remain unknown. The 1.7 A structure of the catalytic domain of NSD1 presented here shows that a regulatory loop adopts a conformation that prevents free access of H3K36 to the bound S-adenosyl-L-methionine. Molecular dynamics simulation and computational docking revealed that this normally inhibitory loop can adopt an active conformation, allowing H3K36 access to the active site, and that the nucleosome may stabilize the active conformation of the regulatory loop. Hence, our study reveals an autoregulatory mechanism of NSD1 and provides insight into the molecular mechanism of the nucleosomal substrate selectivity of this disease-related H3K36 methyltransferase
— id: 138314, year: 2011, vol: 286, page: 8361, stat: Journal Article,

Structure and histone binding properties of the Vps75-Rtt109 chaperone-lysine acetyltransferase complex
Su, Dan; Hu, Qi; Zhou, Hui; Thompson, James R; Xu, Rui-Ming; Zhang, Zhiguo; Mer, Georges
2011 May 6;286(18):15625-15629, Journal of biological chemistry
The histone chaperone Vps75 presents the remarkable property of stimulating the Rtt109-dependent acetylation of several histone H3 lysine residues within (H3-H4)(2) tetramers. To investigate this activation mechanism, we determined x-ray structures of full-length Vps75 in complex with full-length Rtt109 in two crystal forms. Both structures show similar asymmetric assemblies of a Vps75 dimer bound to an Rtt109 monomer. In the Vps75-Rtt109 complexes, the catalytic site of Rtt109 is confined to an enclosed space that can accommodate the N-terminal tail of histone H3 in (H3-H4)(2). Investigation of Vps75-Rtt109-(H3-H4)(2) and Vps75-(H3-H4)(2) complexes by NMR spectroscopy-probed hydrogen/deuterium exchange suggests that Vps75 guides histone H3 in the catalytic enclosure. These findings clarify the basis for the enhanced acetylation of histone H3 tail residues by Vps75-Rtt109
— id: 141095, year: 2011, vol: 286, page: 15625, stat: Journal Article,

KRAS Mutations Are Associated with Specific Morphologic Features in Colon Cancer
Gunal, A; Kenney, B; Hui, P; Kilic, S; Xu, R; Jain, D; Mitchell, K; Robert, M
2010 FEB ;90(11):146A-146A, Laboratory investigation
— id: 109951, year: 2010, vol: 90, page: 146A, stat: Journal Article,

KRAS Mutations Are Associated with Specific Morphologic Features in Colon Cancer
Gunal, A; Kenney, B; Hui, P; Kilie, S; Xu, R; Jain, D; Mitchell, K; Robert, M
2010 FEB ;23(3):146A-146A, Modern pathology
— id: 109932, year: 2010, vol: 23, page: 146A, stat: Journal Article,

The postsynaptic density proteins homer and shank form a polymeric network structure
Hayashi M.; Tang C.; Verpelli C.; Narayanan R.; Stearns M.; Xu R.-M.; Li H.; Sala C.; Hayashi Y.
2010 ;68:e339-e339, Neuroscience research
The postsynaptic density (PSD) is crucial for synaptic functions, but the molecular architecture retaining its structure and components remains elusive. Homer and Shank are among the most abundant scaffolding proteins in the PSD, working synergistically for maturation and enlargement of dendritic spines. Here, we demonstrate that Homer and Shank form a polymeric complex, which could be precipitated by ultracentrifuge. The dynamic light scattering measurement of the complex showed that the size of the complex reaches micrometers, which is large enough to span the size of a dendritic spine. Electron microscope observation revealed amorphous mesh-like matrix structure of the Homer-Shank complex. Crystallographic analysis of the coiled-coil region of Homer revealed a pair of parallel dimeric coiledcoils intercalated in a tail-to-tail fashion to form a tetramer. This indicates the unique configuration of a pair of amino-terminal EVH1 domains at each end of the coiled-coil. The length of the coiled-coil is estimated to be around 45 nm, which is long enough to penetrate the thickness of the PSD, and to connect proteins on plasma membrane and ER. In neurons, the tetramerization is required for structural integrity of the dendritic spines and recruitment of proteins to synapses. We propose that the Homer-Shank complex serves as a structural framework and as an assembly platform for other PSD proteins. Other cellular functions of Homer related to the unique tetrameric coiled-coil structure will be also discussed
— id: 113673, year: 2010, vol: 68, page: e339, stat: Journal Article,

The postsynaptic density proteins homer and shank form a polymeric network structure
Hayashi, M; Tang, CY; Verpelli, C; Narayanan, R; Stearns, M; Xu, RM; Sala, C; Hayashi, Y
2010 JUN 15 ;112(24):110P-110P, Journal of pharmacological sciences
— id: 110115, year: 2010, vol: 112, page: 110P, stat: Journal Article,

Structural basis for methylarginine-dependent recognition of Aubergine by Tudor
Liu, Haiping; Wang, Ju-Yu S; Huang, Ying; Li, Zhizhong; Gong, Weimin; Lehmann, Ruth; Xu, Rui-Ming
2010 Sep 1;24(17):1876-1881, Genes & development
Piwi proteins are modified by symmetric dimethylation of arginine (sDMA), and the methylarginine-dependent interaction with Tudor domain proteins is critical for their functions in germline development. Cocrystal structures of an extended Tudor domain (eTud) of Drosophila Tudor with methylated peptides of Aubergine, a Piwi family protein, reveal that sDMA is recognized by an asparagine-gated aromatic cage. Furthermore, the unexpected Tudor-SN/p100 fold of eTud is important for sensing the position of sDMA. The structural information provides mechanistic insights into sDMA-dependent Piwi-Tudor interaction, and the recognition of sDMA by Tudor domains in general
— id: 113782, year: 2010, vol: 24, page: 1876, stat: Journal Article,

The postsynaptic density proteins Homer and Shank form a polymeric network structure
Hayashi, Mariko Kato; Tang, Chunyan; Verpelli, Chiara; Narayanan, Radhakrishnan; Stearns, Marissa H; Xu, Rui-Ming; Li, Huilin; Sala, Carlo; Hayashi, Yasunori
2009 Apr 3;137(1):159-171, Cell
The postsynaptic density (PSD) is crucial for synaptic functions, but the molecular architecture retaining its structure and components remains elusive. Homer and Shank are among the most abundant scaffolding proteins in the PSD, working synergistically for maturation of dendritic spines. Here, we demonstrate that Homer and Shank, together, form a mesh-like matrix structure. Crystallographic analysis of this region revealed a pair of parallel dimeric coiled coils intercalated in a tail-to-tail fashion to form a tetramer, giving rise to the unique configuration of a pair of N-terminal EVH1 domains at each end of the coiled coil. In neurons, the tetramerization is required for structural integrity of the dendritic spines and recruitment of proteins to synapses. We propose that the Homer-Shank complex serves as a structural framework and as an assembly platform for other PSD proteins
— id: 141101, year: 2009, vol: 137, page: 159, stat: Journal Article,

The target of the NSD family of histone lysine methyltransferases depends on the nature of the substrate
Li, Yan; Trojer, Patrick; Xu, Chong-Feng; Cheung, Peggie; Kuo, Alex; Drury, William J 3rd; Qiao, Qi; Neubert, Thomas A; Xu, Rui-Ming; Gozani, Or; Reinberg, Danny
2009 Dec 4;284(49):34283-34295, Journal of biological chemistry
The NSD (nuclear receptor SET domain-containing) family of histone lysine methyltransferases is a critical participant in chromatin integrity as evidenced by the number of human diseases associated with the aberrant expression of its family members. Yet, the specific targets of these enzymes are not clear, with marked discrepancies being reported in the literature. We demonstrate that NSD2 can exhibit disparate target preferences based on the nature of the substrate provided. The NSD2 complex purified from human cells and recombinant NSD2 both exhibit specific targeting of histone H3 lysine 36 (H3K36) when provided with nucleosome substrates, but histone H4 lysine 44 is the primary target in the case of octamer substrates, irrespective of the histones being native or recombinant. This disparity is negated when NSD2 is presented with octamer targets in conjunction with short single- or double-stranded DNA. Although the octamers cannot form nucleosomes, the target is nonetheless nucleosome-specific as is the product, dimethylated H3K36. This study clarifies in part the previous discrepancies reported with respect to NSD targets. We propose that DNA acts as an allosteric effector of NSD2 such that H3K36 becomes the preferred target
— id: 105498, year: 2009, vol: 284, page: 34283, stat: Journal Article,

An intact unfolded protein response in Trpt1 knockout mice reveals phylogenic divergence in pathways for RNA ligation
Harding, Heather P; Lackey, Jeremy G; Hsu, Hao-Chi; Zhang, Yuhong; Deng, Jing; Xu, Rui-Ming; Damha, Masad J; Ron, David
2008 Feb;14(2):225-232, RNA
Unconventional mRNA splicing by an endoplasmic reticulum stress-inducible endoribonuclease, IRE1, is conserved in all known eukaryotes. It controls the expression of a transcription factor, Hac1p/XBP-1, that regulates gene expression in the unfolded protein response. In yeast, the RNA fragments generated by Ire1p are ligated by tRNA ligase (Trl1p) in a process that leaves a 2'-PO4(2-) at the splice junction, which is subsequently removed by an essential 2'-phosphotransferase, Tpt1p. However, animals, unlike yeast, have two RNA ligation/repair pathways that could potentially rejoin the cleaved Xbp-1 mRNA fragments. We report that inactivation of the Trpt1 gene, encoding the only known mammalian homolog of Tpt1p, eliminates all detectable 2'-phosphotransferase activity from cultured mouse cells but has no measurable effect on spliced Xbp-1 translation. Furthermore, the relative translation rates of tyrosine-rich proteins is unaffected by the Trpt1 genotype, suggesting that the pool of (normally spliced) tRNA(Tyr) is fully functional in the Trpt1-/- mouse cells. These observations argue against the presence of a 2'-PO4(2-) at the splice junction of ligated RNA molecules in Trpt1-/- cells, and suggest that Xbp-1 and tRNA ligation proceed by distinct pathways in yeast and mammals
— id: 76337, year: 2008, vol: 14, page: 225, stat: Journal Article,

Rtt109 acetylates histone H3 lysine 56 and functions in DNA replication
Han, Junhong; Zhou, Hui; Horazdovsky, Bruce; Zhang, Kangling; Xu, Rui-Ming; Zhang, Zhiguo
2007 Feb 2;315(5812):653-655, Science
Acetylation of histone H3 lysine 56 (H3-K56) occurs in S phase, and cells lacking H3-K56 acetylation are sensitive to DNA-damaging agents. However, the histone acetyltransferase (HAT) that catalyzes global H3-K56 acetylation has not been found. Here we show that regulation of Ty1 transposition gene product 109 (Rtt109) is an H3-K56 HAT. Cells lacking Rtt109 or expressing rtt109 mutants with alterations at a conserved aspartate residue lose H3-K56 acetylation and exhibit increased sensitivity toward genotoxic agents, as well as elevated levels of spontaneous chromosome breaks. Thus, Rtt109, which shares no sequence homology with any other known HATs, is a unique HAT that acetylates H3-K56
— id: 141097, year: 2007, vol: 315, page: 653, stat: Journal Article,

Acetylation of lysine 56 of histone H3 catalyzed by RTT109 and regulated by ASF1 is required for replisome integrity
Han, Junhong; Zhou, Hui; Li, Zhizhong; Xu, Rui-Ming; Zhang, Zhiguo
2007 Sep 28;282(39):28587-28596, Journal of biological chemistry
In budding yeast, acetylation of histone H3 lysine 56 (H3-K56) is catalyzed by the Rtt109-Vps75 histone acetyltransferase (HAT) complex, with Rtt109 being the catalytic subunit, and histone chaperone Asf1 is required for this modification. Cells lacking Rtt109 are susceptible to perturbations in DNA replication. However, how Asf1 regulates acetylation of H3-K56 and how loss of H3-K56 acetylation affects DNA replication are unclear. We show that at low concentrations the Rtt109-Vps75 HAT complex acetylates H3-K56 in vitro when H3/H4 is complexed with Asf1, but not H3/H4 tetramers, recapitulating the in vivo requirement of Asf1 for H3-K56 acetylation using recombinant proteins. Moreover, the Rtt109-Vps75 complex interacts with Asf1-H3/H4 but not Asf1. In vivo, the Rtt109-Asf1 interaction is also dependent on the ability of Asf1 to bind H3/H4. Furthermore, the Rtt109 homolog in Schizosaccharomyces pombe (SpRtt109) also displayed an Asf1-dependent H3-K56 HAT activity in vitro. These results indicate that Asf1 regulates H3-K56 acetylation by presenting histones H3 and H4 to Rtt109-Vps575 for acetylation, and this mechanism is likely to be conserved. Finally, we have shown that cells lacking Rtt109 or expressing H3-K56 mutants exhibited significant reduction in the association of three proteins with stalled DNA replication forks and hyper-recombination of replication forks stalled at replication fork barriers of the ribosomal DNA locus compared with wild-type cells. Taken together, these studies provide novel insight into the role of Asf1 in the regulation of H3-K56 acetylation and the function of this modification in DNA replication
— id: 141102, year: 2007, vol: 282, page: 28587, stat: Journal Article,

The Rtt109-Vps75 histone acetyltransferase complex acetylates non-nucleosomal histone H3
Han, Junhong; Zhou, Hui; Li, Zhizhong; Xu, Rui-Ming; Zhang, Zhiguo
2007 May 11;282(19):14158-14164, Journal of biological chemistry
Acetylation of lysine 56 of histone H3 (H3-Lys-56) occurs in S phase and disappears during G(2)/M phase of the cell cycle. However, it is not clear how this modification is regulated during the progression of the cell cycle. We and others have shown that the histone acetyltransferase (HAT) Rtt109 is the primary HAT responsible for acetylating H3-Lys-56 in budding yeast. Here we show that Rtt109 forms a complex with Vps75 and that both recombinant Rtt109-Vps75 complexes and native complexes purified from yeast cells acetylate H3 present in H3/H4/H2A/H2B core histones but not other histones. In addition, both recombinant and native Rtt109-Vps75 HAT complexes exhibited no detectable activity toward nucleosomal H3, suggesting that H3-Lys-56 acetylation is at least in part regulated by the inability of Rtt109-Vps75 complexes to acetylate nucleosomal H3 during G(2)/M phase of the cell cycle. Further, Rtt109 bound mutant H3/H4 tetramers composed of histones lacking their N-terminal tail domains less efficiently than wild-type H3/H4 tetramers, and Rtt109-Vps75 complexes displayed reduced HAT activity toward these mutant H3/H4 tetramers. Thus, the N termini of H3/H4 tetramers are required for efficient acetylation of H3 by the Rtt109-Vps75 complex. Taken together, these studies provide insights into how H3-Lys-56 acetylation is regulated during the cell cycle
— id: 141103, year: 2007, vol: 282, page: 14158, stat: Journal Article,

Structural basis for the function of DCN-1 in protein Neddylation
Yang, Xiaoyu; Zhou, Jie; Sun, Lei; Wei, Zhiyi; Gao, Jianying; Gong, Weimin; Xu, Rui-Ming; Rao, Zihe; Liu, Yingfang
2007 Aug 24;282(34):24490-24494, Journal of biological chemistry
Covalent modification by Nedd8 (neddylation) stimulates the ubiquitin-protein isopeptide ligase (E3) activities of Cullins. DCN-1, an evolutionarily conserved protein, promotes neddylation of Cullins in vivo, binds directly to Nedd8, and associates with Cdc53 in the budding yeast Saccharomyces cerevisiae. The 1.9A resolution structure of yeast DCN-1 shows that the region encompassing residues 66-269 has a rectangular parallelepiped-like all alpha-helical structures, consisting of an EF-hand motif N-terminal domain and a closely juxtaposed C-terminal domain with six alpha-helices. The EF-hand motif structure is highly similar to that of the c-Cbl ubiquitin E3 ligase. We also demonstrate that DCN-1 directly binds to Rbx-1, a factor important for protein neddylation. The structural and biochemical results are consistent with the role of DCN-1 as a scaffold protein in a multisubunit neddylation E3 ligase complex
— id: 141100, year: 2007, vol: 282, page: 24490, stat: Journal Article,

Structure of the Mycobacterium tuberculosis proteasome and mechanism of inhibition by a peptidyl boronate
Hu, Guiqing; Lin, Gang; Wang, Ming; Dick, Lawrence; Xu, Rui-Ming; Nathan, Carl; Li, Huilin
2006 Mar;59(5):1417-1428, Molecular microbiology
Mycobacterium tuberculosis (Mtb) has the remarkable ability to resist killing by human macrophages. The 750 kDa proteasome, not available in most eubacteria except Actinomycetes, appears to contribute to Mtb's resistance. The crystal structure of the Mtb proteasome at 3.0 A resolution reveals a substrate-binding pocket with composite features of the distinct beta1, beta2 and beta5 substrate binding sites of eukaryotic proteasomes, accounting for the broad specificity of the Mtb proteasome towards oligopeptides described in the companion article [Lin et al. (2006), Mol Microbiol doi:10.1111/j.1365-2958.2005.05035.x]. The substrate entrance at the end of the cylindrical proteasome appears open in the crystal structure due to partial disorder of the alpha-subunit N-terminal residues. However, cryo-electron microscopy of the core particle reveals a closed end, compatible with the density observed in negative-staining electron microscopy that depended on the presence of the N-terminal octapetides of the alpha-subunits in the companion article, suggesting that the Mtb proteasome has a gated structure. We determine for the first time the proteasomal inhibition mechanism of the dipeptidyl boronate N-(4-morpholine)carbonyl-beta-(1-naphthyl)-l-alanine-l-leucine boronic acid (MLN-273), an analogue of the antimyeloma drug bortezomib. The structure improves prospects for designing Mtb-specific proteasomal inhibitors as a novel approach to chemotherapy of tuberculosis.
— id: 62660, year: 2006, vol: 59, page: 1417, stat: Journal Article,

Recognition of histone H3 lysine-4 methylation by the double tudor domain of JMJD2A
Huang, Ying; Fang, Jia; Bedford, Mark T; Zhang, Yi; Xu, Rui-Ming
2006 May 5;312(5774):748-751, Science
Biological responses to histone methylation critically depend on the faithful readout and transduction of the methyl-lysine signal by 'effector' proteins, yet our understanding of methyl-lysine recognition has so far been limited to the study of histone binding by chromodomain and WD40-repeat proteins. The double tudor domain of JMJD2A, a Jmjc domain-containing histone demethylase, binds methylated histone H3-K4 and H4-K20. We found that the double tudor domain has an interdigitated structure, and the unusual fold is required for its ability to bind methylated histone tails. The cocrystal structure of the JMJD2A double tudor domain with a trimethylated H3-K4 peptide reveals that the trimethyl-K4 is bound in a cage of three aromatic residues, two of which are from the tudor-2 motif, whereas the binding specificity is determined by side-chain interactions involving amino acids from the tudor-1 motif. Our study provides mechanistic insights into recognition of methylated histone tails by tudor domains and reveals the structural intricacy of methyl-lysine recognition by two closely spaced effector domains
— id: 141098, year: 2006, vol: 312, page: 748, stat: Journal Article,

Structure of a Bmi-1-Ring1B polycomb group ubiquitin ligase complex
Li, Zhizhong; Cao, Ru; Wang, Ming; Myers, Michael P; Zhang, Yi; Xu, Rui-Ming
2006 Jul 21;281(29):20643-20649, Journal of biological chemistry
Polycomb group proteins Bmi-1 and Ring1B are core subunits of the PRC1 complex, which plays important roles in the regulation of Hox gene expression, X-chromosome inactivation, tumorigenesis, and stem cell self-renewal. The RING finger protein Ring1B is an E3 ligase that participates in the ubiquitination of lysine 119 of histone H2A, and the binding of Bmi-1 stimulates the E3 ligase activity. We have mapped the regions of Bmi-1 and Ring1B required for efficient ubiquitin transfer and determined a 2.5-A structure of the Bmi-1-Ring1B core domain complex. The structure reveals that Ring1B 'hugs' Bmi-1 through extensive RING domain contacts and its N-terminal tail wraps around Bmi-1. The two regions of interaction have a synergistic effect on the E3 ligase activity. Our analyses suggest a model where the Bmi-1-Ring1B complex stabilizes the interaction between the E2 enzyme and the nucleosomal substrate to allow efficient ubiquitin transfer
— id: 141099, year: 2006, vol: 281, page: 20643, stat: Journal Article,

Structural basis for origin recognition complex 1 protein-silence information regulator 1 protein interaction in epigenetic silencing
Hsu, Hao-Chi; Stillman, Bruce; Xu, Rui-Ming
2005 Jun 14;102(24):8519-8524, Proceedings of the National Academy of Sciences of the United States of America
The interaction between silence information regulator 1 protein (Sir1p) and origin recognition complex 1 protein (Orc1p), the largest subunit of the origin recognition complex, plays an important role in the establishment of transcriptional silencing at the cryptic mating-type gene loci in Saccharomyces cerevisiae. Sir1p binds the N-terminal region of Orc1p encompassing a Bromo-adjacent homology (BAH) domain found in various chromatin-associated proteins. To understand the molecular mechanism of Sir protein recruitment, we have determined a 2.5-A cocrystal structure of the N-terminal domain of Orc1p in complex with the Orc1p-interacting domain of Sir1p. The structure reveals that Sir1p Orc1p-interacting domain has a bilobal structure: an alpha/beta N-terminal lobe and a C-terminal lobe resembling the Tudor domain royal family fold. The N-terminal lobe of Sir1p binds in a shallow groove between a helical subdomain and the BAH domain of Orc1p. The structure provides a mechanistic understanding of Orc1p-Sir1p interaction specificity, as well as insights into protein-protein interactions involving BAH domains in general.
— id: 62661, year: 2005, vol: 102, page: 8519, stat: Journal Article,

RNA recognition motif 2 directs the recruitment of SF2/ASF to nuclear stress bodies
Chiodi, Ilaria; Corioni, Margherita; Giordano, Manuela; Valgardsdottir, Rut; Ghigna, Claudia; Cobianchi, Fabio; Xu, Rui-Ming; Riva, Silvano; Biamonti, Giuseppe
2004 ;32(14):4127-4136, Nucleic acids research
Heat shock induces the transcriptional activation of large heterochromatic regions of the human genome composed of arrays of satellite III DNA repeats. A number of RNA-processing factors, among them splicing factor SF2/ASF, associate with these transcription factors giving rise to nuclear stress bodies (nSBs). Here, we show that the recruitment of SF2/ASF to these structures is mediated by its second RNA recognition motif. Amino acid substitutions in the first alpha-helix of this domain, but not in the beta-strand regions, abrogate the association with nSBs. The same mutations drastically affect the in vivo activity of SF2/ASF in the alternative splicing of adenoviral E1A transcripts. Sequence analysis identifies four putative high-affinity binding sites for SF2/ASF in the transcribed strand of the satellite III DNA. We have verified by gel mobility shift assays that the second RNA-binding domain of SF2/ASF binds at least one of these sites. Our analysis suggests that the recruitment of SF2/ASF to nSBs is mediated by a direct interaction with satellite III transcripts and points to the second RNA-binding domain of the protein as the major determinant of this interaction.
— id: 62664, year: 2004, vol: 32, page: 4127, stat: Journal Article,

Crystal structure of the human ATP-dependent splicing and export factor UAP56
Shi, Hang; Cordin, Olivier; Minder, C Michael; Linder, Patrick; Xu, Rui-Ming
2004 Dec 21;101(51):17628-17633, Proceedings of the National Academy of Sciences of the United States of America
Pre-mRNA splicing requires the function of a number of RNA-dependent ATPases/helicases, yet no three-dimensional structure of any spliceosomal ATPases/helicases is known. The highly conserved DECD-box protein UAP56/Sub2 is an essential splicing factor that is also important for mRNA export. The expected ATPase/helicase activity appears to be essential for the UAP56/Sub2 functions. Here, we show that purified human UAP56 is an active RNA-dependent ATPase, and we also report the crystal structures of UAP56 alone and in complex with ADP, as well as a DECD to DEAD mutant. The structures reveal a unique spatial arrangement of the two conserved helicase domains, and ADP-binding induces significant conformational changes of key residues in the ATP-binding pocket. Our structural analyses suggest a specific protein-RNA displacement model of UAP56/Sub2. The detailed structural information provides important mechanistic insights into the splicing function of UAP56/Sub2. The structures also will be useful for the analysis of other spliceosomal DExD-box ATPases/helicases.
— id: 62663, year: 2004, vol: 101, page: 17628, stat: Journal Article,

Set2-catalyzed methylation of histone H3 represses basal expression of GAL4 in Saccharomyces cerevisiae
Landry, Joseph; Sutton, Ann; Hesman, Tina; Min, Jinrong; Xu, Rui-Ming; Johnston, Mark; Sternglanz, Rolf
2003 Sep;23(17):5972-5978, Molecular & cellular biology
Recent work has shown that histone methylation is an important regulator of transcription. While much is known about the roles of histone methyltransferases (HMTs) in the establishment of heterochromatin, little is known of their roles in the regulation of actively transcribed genes. We describe an in vivo role of the Saccharomyces cerevisiae HMT, Set2. We identified SET2 as a gene necessary for repression of GAL4 basal expression and show that the evolutionarily conserved SACI, SACII, and SET domains of Set2 are necessary for this repression. We confirm that Set2 catalyzes methylation of lysine 36 on the N-terminal tail of histone H3. Conversion of lysine 36 to an unmethylatable arginine causes a decrease in the repression of GAL4 transcription, as does a Delta set2 mutation. We further show that lysine 36 of histone H3 at GAL4 is methylated and that this methylation is dependent upon the presence of SET2.
— id: 62665, year: 2003, vol: 23, page: 5972, stat: Journal Article,

Structure of the catalytic domain of human DOT1L, a non-SET domain nucleosomal histone methyltransferase
Min, Jinrong; Feng, Qin; Li, Zhizhong; Zhang, Yi; Xu, Rui-Ming
2003 Mar 7;112(5):711-723, Cell
Dot1 is an evolutionarily conserved histone methyltransferase that methylates lysine-79 of histone H3 in the core domain. Unlike other histone methyltransferases, Dot1 does not contain a SET domain, and it specifically methylates nucleosomal histone H3. We have solved a 2.5 A resolution structure of the catalytic domain of human Dot1, hDOT1L, in complex with S-adenosyl-L-methionine (SAM). The structure reveals a unique organization of a mainly alpha-helical N-terminal domain and a central open alpha/beta structure, an active site consisting of a SAM binding pocket, and a potential lysine binding channel. We also show that a flexible, positively charged region at the C terminus of the catalytic domain is critical for nucleosome binding and enzymatic activity. These structural and biochemical analyses, combined with molecular modeling, provide mechanistic insights into the catalytic mechanism and nucleosomal specificity of Dot1 proteins.
— id: 62669, year: 2003, vol: 112, page: 711, stat: Journal Article,

Structural basis for specific binding of Polycomb chromodomain to histone H3 methylated at Lys 27
Min, Jinrong; Zhang, Yi; Xu, Rui-Ming
2003 Aug 1;17(15):1823-1828, Genes & development
The chromodomain of Drosophila Polycomb protein is essential for maintaining the silencing state of homeotic genes during development. Recent studies suggest that Polycomb mediates the assembly of repressive higher-order chromatin structures in conjunction with the methylation of Lys 27 of histone H3 by a Polycomb group repressor complex. A similar mechanism in heterochromatin assembly is mediated by HP1, a chromodomain protein that binds to histone H3 methylated at Lys 9. To understand the molecular mechanism of the methyl-Lys 27 histone code recognition, we have determined a 1.4-A-resolution structure of the chromodomain of Polycomb in complex with a histone H3 peptide trimethylated at Lys 27. The structure reveals a conserved mode of methyl-lysine binding and identifies Polycomb-specific interactions with histone H3. The structure also reveals a dPC dimer in the crystal lattice that is mediated by residues specifically conserved in the Polycomb family of chromodomains. The dimerization of dPC can effectively account for the histone-binding specificity and provides new mechanistic insights into the function of Polycomb. We propose that self-association is functionally important for Polycomb.
— id: 62666, year: 2003, vol: 17, page: 1823, stat: Journal Article,

Crystal structure of the Drosophila Mago nashi-Y14 complex
Shi, Hang; Xu, Rui-Ming
2003 Apr 15;17(8):971-976, Genes & development
Pre-mRNA splicing is essential for generating mature mRNA and is also important for subsequent mRNA export and quality control. The splicing history is imprinted on spliced mRNA through the deposition of a splicing-dependent multiprotein complex, the exon junction complex (EJC), at approximately 20 nucleotides upstream of exon-exon junctions. The EJC is a dynamic structure containing proteins functioning in the nuclear export and nonsense-mediated decay of spliced mRNAs. Mago nashi (Mago) and Y14 are core components of the EJC, and they form a stable heterodimer that strongly associates with spliced mRNA. Here we report a 1.85 A-resolution structure of the Drosophila Mago-Y14 complex. Surprisingly, the structure shows that the canonical RNA-binding surface of the Y14 RNA recognition motif (RRM) is involved in extensive protein-protein interactions with Mago. This unexpected finding provides important insights for understanding the molecular mechanisms of EJC assembly and RRM-mediated protein-protein interactions.
— id: 62668, year: 2003, vol: 17, page: 971, stat: Journal Article,

A pivotal role of the coiled coil of Sir4
Xu, Rui-Ming
2003 Jun;11(6):608-609, Structure
The C terminus of Sir4 forms a coiled-coil structure. The coiled-coil domain is responsible for the dimerization of Sir4 and contains the binding site of Sir3. Structural and biochemical analyses of the Sir4 coiled-coil domain provide important insights into the molecular mechanisms of Sir3-Sir4 interaction and the assembly of a ternary Sir2/Sir3/Sir4 complex that are essential for epigenetic control of gene expression in S. cerevisiae.
— id: 62667, year: 2003, vol: 11, page: 608, stat: Journal Article,

Structure of the SET domain histone lysine methyltransferase Clr4
Min, Jinrong; Zhang, Xing; Cheng, Xiaodong; Grewal, Shiv I S; Xu, Rui-Ming
2002 Nov;9(11):828-832, Nature structural biology
Methylation of histone H3 lysine 9 is an important component of the 'histone code' for heterochromatic gene silencing. The SET domain-containing Clr4 protein, a close relative of Su(var)3-9 proteins in higher eukaryotes, specifically methylates lysine 9 of histone H3 and is essential for silencing in Schizosaccharomyces pombe. Here we report the 2.3 A resolution crystal structure of the catalytic domain of Clr4. The structure reveals an overall fold rich in beta-strands, a potential active site consisting of a SAM-binding pocket, and a connected groove that could accommodate the binding of the N-terminal tail of histone H3. The pre-SET motif contains a triangular zinc cluster coordinated by nine cysteines distant from the active site, whereas the post-SET region is largely flexible but proximal to the active site. The structure provides insights into the architecture of SET domain histone methyltransferases and establishes a paradigm for further characterization of the Clr4 family of epigenetic regulators.
— id: 62670, year: 2002, vol: 9, page: 828, stat: Journal Article,

Ubiquitination of histone H2B by Rad6 is required for efficient Dot1-mediated methylation of histone H3 lysine 79
Ng, Huck Hui; Xu, Rui-Ming; Zhang, Yi; Struhl, Kevin
2002 Sep 20;277(38):34655-34657, Journal of biological chemistry
Dot1 is a non-SET domain protein that methylates histone H3 at lysine 79, a surface-exposed residue that lies within the globular domain. In the context of a nucleosome, H3 lysine 79 is located in close proximity with lysine 123 of histone H2B, a major site for ubiquitination by Rad6. Here we show that Rad6-mediated ubiquitination of H2B lysine 123 is important for efficient methylation of lysine 79, but not lysine 36, of histone H3. In contrast, lysine 79 methylation of H3 is not required for ubiquitination of H2B. Our study provides a new example of trans-histone regulation between modifications on different histones. In addition, it suggests that Rad6 affects telomeric silencing, at least in part, by influencing methylation of histone H3.
— id: 62672, year: 2002, vol: 277, page: 34655, stat: Journal Article,

Structure and function of the BAH-containing domain of Orc1p in epigenetic silencing
Zhang, Zhiguo; Hayashi, Mariko K; Merkel, Olaf; Stillman, Bruce; Xu, Rui-Ming
2002 Sep 2;21(17):4600-4611, EMBO journal
The N-terminal domain of the largest subunit of the Saccharomyces cerevisiae origin recognition complex (Orc1p) functions in transcriptional silencing and contains a bromo-adjacent homology (BAH) domain found in some chromatin-associated proteins including Sir3p. The 2.2 A crystal structure of the N-terminal domain of Orc1p revealed a BAH core and a non-conserved helical sub-domain. Mutational analyses demonstrated that the helical sub-domain was necessary and sufficient to bind Sir1p, and critical for targeting Sir1p primarily to the cis-acting E silencers at the HMR and HML silent chromatin domains. In the absence of the BAH domain, approximately 14-20% of cells in a population were silenced at the HML locus. Moreover, the distributions of the Sir2p, Sir3p and Sir4p proteins, while normal, were at levels lower than found in wild-type cells. Thus, in the absence of the Orc1p BAH domain, HML resembled silencing of genes adjacent to telomeres. These data are consistent with the view that the Orc1p-Sir1p interaction at the E silencers ensures stable inheritance of pre-established Sir2p, Sir3p and Sir4p complexes at the silent mating type loci.
— id: 62671, year: 2002, vol: 21, page: 4600, stat: Journal Article,