A Conserved SET Domain Methyltransferase, Set11, Modifies Ribosomal Protein Rpl12 in Fission Yeast

Sadaie, Mahito; Shinmyozu, Kaori; Nakayama, Jun‐ichi

doi:10.1074/jbc.m709429200

Cited by 39 publications

(41 citation statements)

References 44 publications

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“…By 2006, our laboratory had found evidence for five added methyl groups on the large ribosomal subunit yeast protein Rpl12ab near the N-terminus; modification at Lys-3 by a SET-domain methyltransferase designated Rkm2 and by a yet unidentified enzyme at Lys-10 (44). However, these results were called into question by reports on the modifications of the orthologs of Rpl12ab in Arabidopsis thaliana (51) and Schizosaccharomyces pombe (52); both reports suggested that our mass spectrometry data was more consistent with the dimethylation of the N-terminal proline residue and the trimethylation of Lys-3 in line with the similar N-terminal modification of the Arabidopsis and S. pombe proteins. Further analysis in our laboratory (45) confirmed the N-terminal modification and the major methylation of Lys-3 as opposed to Lys-10 (53).…”

Section: Protein N-terminal Methyltransferasesmentioning

confidence: 57%

The ribosome: A hot spot for the identification of new types of protein methyltransferases

Clarke¹

2018

Journal of Biological Chemistry

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Cellular physiology depends on the alteration of protein structures by covalent modification reactions. Using a combination of bioinformatic, genetic, biochemical, and mass spectrometric approaches, it has been possible to probe ribosomal proteins from the yeast Saccharomyces cerevisiae for posttranslationally methylated amino acid residues and for the enzymes that catalyze these modifications. These efforts have resulted in the identification and characterization of the first protein histidine methyltransferase, the first N-terminal protein methyltransferase, two unusual types of protein arginine methyltransferases, and a new type of cysteine methylation. Two of these enzymes may modify their substrates during ribosomal assembly because the final methylated histidine and arginine residues are buried deep within the ribosome with contacts only with RNA. Two of these modifications occur broadly in eukaryotes, including humans, while the others demonstrate a more limited phylogenetic range. Analysis of strains where the methyltransferase genes are deleted has given insight into the physiological roles of these modifications. These reactions described here add diversity to the modifications that generate the typical methylated lysine and arginine residues previously described in histones and other proteins. Regulation of biological function by protein methylation reactionsIt is more and more apparent just how much of biological function is dependent upon posttranslational modifications (1). The human genome encodes some 900 enzymes catalyzing just protein phosphorylation or ubiquitination (2,3). However, it is now clear that methyl groups can stand beside phosphate groups and ubiquitin as major players in controlling the physiological functions of proteins. We are beginning to understand how the much greater diversity of protein methylation reactions can give rise to a greater diversity of function (1,4-8). We are also learning the importance of crosstalk between protein and DNA methylation reactions (9) and among protein methylation, phosphorylation, acetylation, and ubiquitination reactions (10-12). Finally, we are discovering how alterations in protein methylation pathways can lead to human pathology, particularly in cancer (13-15).The collection of over sixty human protein arginine and lysine methyltransferases that leave histone "marks" recognized by reader proteins to guide gene expression are perhaps the poster children for the importance of protein methyltransferases (16-17). However, recent work has emphasized the importance of methylating non-histone substrates at these residues, particularly ribosomal proteins, translation factors, and transcription factors in a wide variety of organisms (7,8,15,18,19). Furthermore, the methylation of lysine and arginine residues represents just the tip of the iceberg in protein methylation -modifications have also been established at histidine, modified histidine (diphthamide), glutamate, glutamine, asparagine, Lisoaspartate, D-aspartate, cysteine, isoprenylcysteine, me...

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Section: Protein N-terminal Methyltransferasesmentioning

confidence: 57%

The ribosome: A hot spot for the identification of new types of protein methyltransferases

Clarke¹

2018

Journal of Biological Chemistry

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“…The in vitro methyltransferase assay and the chromatographic fractionation of S. pombe nuclear extracts were performed as described previously (34).…”

Section: Methodsmentioning

confidence: 99%

“…To analyze the localization of green fluorescent protein-fused Rpl42, wild-type and ⌬set13 mutant S. pombe cells that had integrated the pDUAL-GFH1-rpl42 construct were grown and treated as described above. Microscopic images were captured on a Zeiss Two-dimensional Electrophoretic Analysis of Proteins-The two-dimensional gel analysis of methylated proteins was performed as described previously (34).…”

Section: Analysis Of Methylated Peptide By Nano-liquid Chromatographymentioning

confidence: 99%

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Methylation of Ribosomal Protein L42 Regulates Ribosomal Function and Stress-adapted Cell Growth

Shirai

Sadaie

Shinmyozu³

et al. 2010

Journal of Biological Chemistry

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Lysine methylation is one of the most common protein modifications. Although lysine methylation of histones has been extensively studied and linked to gene regulation, that of nonhistone proteins remains incompletely understood. Here, we show a novel regulatory role of ribosomal protein methylation. Using an in vitro methyltransferase assay, we found that Schizosaccharomyces pombe Set13, a SET domain protein encoded by SPAC688.14, specifically methylates lysine 55 of ribosomal protein L42 (Rpl42). Mass spectrometric analysis revealed that endogenous Rpl42 is monomethylated at lysine 55 in wild-type S. pombe cells and that the methylation is lost in ⌬set13 mutant cells. ⌬set13 and Rpl42 methylation-deficient mutant S. pombe cells showed higher cycloheximide sensitivity and defects in stress-responsive growth control compared with wild type. Genetic analyses suggested that the abnormal growth phenotype was distinct from the conserved stress-responsive pathway that modulates translation initiation. Furthermore, the Rpl42 methylation-deficient mutant cells showed a reduced ability to survive after entering stationary phase. These results suggest that Rpl42 methylation plays direct roles in ribosomal function and cell proliferation control independently of the general stress-response pathway.Post-translational modifications regulate protein structure and function, and one of the most common is lysine methylation (1). Although lysine methylation does not change the overall charge of the side chain, it can influence the activity of protein by providing a novel interface for its interactions with other molecules. Most protein lysine methylations, with a few exceptions, are catalyzed by the SET domain-containing protein (SET protein) family (2). The SET domain was originally identified in three Drosophila proteins, Su(var)3-9, Enhancer of zest, and Trithorax, and was later demonstrated to be the responsible domain for histone-lysine methyltransferase function (3). The characterization of these enzymes has revealed the roles of histone lysine methylation in many different biological processes, including higher order chromatin assembly and transcriptional regulation (4). Recent studies have shown that histone lysine methylation is directly reversed by several histone demethylase families (5).Lysine methylation has also been identified in non-histone proteins that include ribulose 1,5-bisphosphate carboxylase/ oxygenase in plants (6), cytochrome c in yeast (7), mammalian TAF10 and p53 (8 -10), and ribosomal proteins in a diverse range of species (11). The methylation of ribosomal proteins has been observed in both prokaryotes and eukaryotes. In the budding yeast Saccharomyces cerevisiae, a combination of in vivo labeling and direct mass spectrometric analysis of the ribosomal proteins revealed that six of them, Rpl1, Rpl3, Rpl12, Rpl23, Rpl42, and Rpl43, are post-translationally methylated (12, 13). By analyzing the methylation state of S. cerevisiae mutant strains with deletions in candidate SET domain-containing genes, tw...

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“…After silver staining with SilverQuest TM Silver Staining Kit (Invitrogen), the peptide bands were excised from the gel and subjected to ingel reduction with 10 mM DTT, alkylation with 55 mM iodoacetamide, and digestion with 10 μg/ml modified trypsin (Promega) at 37°C for 16 hours. After in-gel digestion, the collected peptides were subjected to mass spectrometry analysis as described previously (Sadaie et al, 2008).…”

Section: Construction Of Gene Disruption Strainsmentioning

confidence: 99%

Fub1p, a novel protein isolated by boundary screening, binds the proteasome complex

et al. 2011

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Silenced chromatin domains are restricted to specific regions. Eukaryotic chromosomes are organized into discrete domains delimited by domain boundaries. From approximately 6,000 genes in Saccharomyces cerevisiae, we previously isolated 55 boundary genes. In this study, we focus on the molecular function of one of boundary genes, YCR076C/FUB1 (function of boundary), whose function has not been clearly defined in vivo. Biochemical analysis of Fub1p revealed that it interacted with multiple subunits of the 20S proteasome core particle (20S CP). To further clarify the functional link between Fub1p and proteasome, several proteasome mutants were analyzed. Although only 20S CP subunits were isolated as Fub1p interactors, a genetic interaction was also observed for component of 19S regulatory particle (19S RP) suggesting involvement of Fub1p with the whole proteasome. We also analyzed the mechanism of boundary establishment by using proteasome composition factor-deficient strains. Deletion of pre9 and ump1, whose products have effects on the 20S CP, resulted in a decrease in boundary function. Domain analyses of Fub1p identified a minimum functional domain in the C terminus that was essential for boundary establishment and showed a limited sequence homology to the human PSMF1, which is known to inhibit proteasome activity. Finally, boundary assay showed that human PSMF1 also exhibited boundary establishment activity in yeast. Our results defined the functional correlation between Fub1p and PSMF1.

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A Conserved SET Domain Methyltransferase, Set11, Modifies Ribosomal Protein Rpl12 in Fission Yeast

Cited by 39 publications

References 44 publications

The ribosome: A hot spot for the identification of new types of protein methyltransferases

The ribosome: A hot spot for the identification of new types of protein methyltransferases

Methylation of Ribosomal Protein L42 Regulates Ribosomal Function and Stress-adapted Cell Growth

Fub1p, a novel protein isolated by boundary screening, binds the proteasome complex

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