Molecular Analysis ofMETTL1,a Novel Human Methyltransferase-like Gene with a High Degree of Phylogenetic Conservation

Bahr, A.; Hankeln, Thomas; Fiedler, Thomas; Hegemann, Johannes H.; Schmidt, Erwin R.

doi:10.1006/geno.1999.5780

Cited by 32 publications

(34 citation statements)

References 8 publications

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“…This phenotype underscores the importance of these proteins in survival, as implied by the widespread evolutionary conservation of putative orthologs of Trm8p and Trm82p (Bahr et al 1999;Michaud et al 2000;Alexandrov et al 2002). The finding of nearly identical phenotypes of trm8-⌬ and trm82-⌬ mutants extends our previous genetic and biochemical data implicating TRM8 and TRM82 in the same process, as indicated by their mutual participation in tRNA m 7 G modification in vitro and in vivo, and by their presence in the same complex (Alexandrov et al 2002).…”

Section: Discussionsupporting

confidence: 80%

“…Neither Trm8p nor Trm82p is significantly related to yeast Abd1p, which catalyzes m 7 G formation during capping of mRNAs (Mao et al 1995), other than within the S-adenosylmethionine (SAM)-binding domain of Trm8p. Trm8p is highly conserved in eukaryotes and bacteria, extending from humans to Mycoplasma genitalium, with less that 500 ORFs (Bahr et al 1999), and Trm82p orthologs are present in the majority of sequenced eukaryotes. Nevertheless, deletion of TRM8 or TRM82 results in no obvious growth defect under standard laboratory conditions (Alexandrov et al 2002), raising the question of the role of these proteins in these organisms.…”

Section: Introductionmentioning

confidence: 99%

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tRNA m⁷G methyltransferase Trm8p/Trm82p: Evidence linking activity to a growth phenotype and implicating Trm82p in maintaining levels of active Trm8p

Alexandrov¹,

Grayhack²,

Phizicky³

2005

RNA

113

115

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We show that Saccharomyces cerevisiae strains lacking Trm8p/Trm82p tRNA m 7 G methyltransferase are temperature-sensitive in synthetic media containing glycerol. Bacterial TRM8 orthologs complement the growth defect of trm8-⌬, trm82-⌬, and trm8-⌬ trm82-⌬ double mutants, suggesting that bacteria employ a single subunit for Trm8p/Trm82p function. The growth phenotype of trm8 mutants correlates with lack of tRNA m 7 G methyltransferase activity in vitro and in vivo, based on analysis of 10 mutant alleles of trm8 and bacterial orthologs, and suggests that m 7 G modification is the cellular function important for growth. Initial examination of the roles of the yeast subunits shows that Trm8p has most of the functions required to effect m 7 G modification, and that a major role of Trm82p is to maintain cellular levels of Trm8p. Trm8p efficiently cross-links to pretRNA Phe in vitro in the presence or absence of Trm82p, in addition to its known residual tRNA m 7 G modification activity and its SAM-binding domain. Surprisingly, the levels of Trm8p, but not its mRNA, are severely reduced in a trm82-⌬ strain. Although Trm8p can be produced in the absence of Trm82p by deliberate overproduction, the resulting protein is inactive, suggesting that a second role of Trm82p is to stabilize Trm8p in an active conformation.

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Section: Discussionsupporting

confidence: 80%

Section: Introductionmentioning

confidence: 99%

tRNA m⁷G methyltransferase Trm8p/Trm82p: Evidence linking activity to a growth phenotype and implicating Trm82p in maintaining levels of active Trm8p

Alexandrov¹,

Grayhack²,

Phizicky³

2005

RNA

113

115

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“…HMETTL1 has two predicted isoforms from two transcript variants, while hWD4 has two transcript variants that only differ in their 3¢ untranslated regions. HMETTL1 was originally suspected to be an RNA methylase of some sort (Bahr et al, 1999), while hWDR4 was originally found in a search for Down Syndrome genetic factors at chromosome 21 (Michaud et al, 2000). The WDR4 gene is located in the Down Syndrome chromosome deleted region (21q22.3), so WDR4 remains a potential candidate for Down Syndrome; nevertheless, further work is required to make any definite connection between the protein and the disease.…”

Section: Sctrm8/82mentioning

confidence: 99%

Transfer RNA Methytransferases and Their Corresponding Modifications in Budding Yeast and Humans: Activities, Predications, and Potential Roles in Human Health

Towns

Begley

2012

DNA and Cell Biology

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Throughout the kingdoms of life, transfer RNA (tRNA) undergoes over 100 enzyme-catalyzed, methyl-based modifications. Although a majority of the methylations are conserved from bacteria to mammals, the functions of a number of these modifications are unknown. Many of the proteins responsible for tRNA methylation, named tRNA methyltransferases (Trms), have been characterized in Saccharomyces cerevisiae. In contrast, only a few human Trms have been characterized. A BLAST search for human homologs of each S. cerevisiae Trm revealed a total of 34 human proteins matching our search criteria for an S. cerevisiae Trm homolog candidate. We have compiled a database cataloging basic information about each human and yeast Trm. Every S. cerevisiae Trm has at least one human homolog, while several Trms have multiple candidates. A search of cancer cell versus normal cell mRNA expression studies submitted to Oncomine found that 30 of the homolog genes display a significant change in mRNA expression levels in at least one data set. While 6 of the 34 human homolog candidates have confirmed tRNA methylation activity, the other candidates remain uncharacterized. We believe that our database will serve as a resource for investigating the role of human Trms in cellular stress signaling.

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“…used for galactose-induced expression for immunopurification experiments+ ORFs were expressed under P GAL control using the BamHIEcoRI GAL1-GAL10 promoter region from pBM258 (Johnston & Davis, 1984), which was ligated into the URA3 CEN vector yCPlac33 and LEU2 CEN vector yCPlac111 (Gietz & Sugino, 1988) to produce vectors pAVA0040 and pAVA0042, respectively+ ORFs YDL201w and YDR165w were PCR-amplified from yeast genomic DNA with appropriate primer pairs (201_F2,201_R,and 165_R, and ORFs of human proteins METTL1 (Bahr et al+, 1999) and WDR4 (Michaud et al+, 2000) were PCR-amplified from I+M+A+G+E+ Consortium clones 3163932 and 4080041 using appropriate primer pairs (METTL1_Fwd: 59-ATCATGCCATGGGATCCACAAT GGCAGCCGAGACTCGGAAC-39; METTL1_Rev: 59-CAT CGCGGATCCGTCGACTCAGTGACCAGGCAGGCTGG-39; WDR4_Fwd: 59-ATCATGCCATGGGATCCACAATGGCGG GCTCTGTGGGAC-39; and WDR4_Rev: 59-CATCGCGGA TCCGTCGACTCAGCAACTTAGCGTCGCCTCC-39)+ ORFs YDL201w and YDR165w were digested with BamHI and Pst I, and ORFs of METTL1 and WDR4 were digested with BamHI and Sal I+ Digested ORFs were ligated into vectors pAVA0040 and pAVA0042 to place the genes under P GAL1 control+…”

mentioning

confidence: 99%

Two proteins that form a complex are required for 7-methylguanosine modification of yeast tRNA

Alexandrov

Martzen

Phizicky

2002

RNA

288

278

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ABSTRACT7-methylguanosine (m 7 G) modification of tRNA occurs widely in eukaryotes and bacteria, is nearly always found at position 46, and is one of the few modifications that confers a positive charge to the base. Screening of a Saccharomyces cerevisiae genomic library of purified GST-ORF fusion proteins reveals two previously uncharacterized proteins that copurify with m 7 G methyltransferase activity on pre-tRNA Phe . ORF YDL201w encodes Trm8, a protein that is highly conserved in prokaryotes and eukaryotes and that contains an S-adenosylmethionine binding domain. ORF YDR165w encodes Trm82, a less highly conserved protein containing putative WD40 repeats, which are often implicated in macromolecular interactions. Neither protein has significant sequence similarity to yeast Abd1, which catalyzes m 7 G modification of the 59 cap of mRNA, other than the methyltransferase motif shared by Trm8 and Abd1. Several lines of evidence indicate that both Trm8 and Trm82 proteins are required for tRNA m 7 G-methyltransferase activity: Extracts derived from strains lacking either gene have undetectable m 7 G methyltransferase activity, RNA from strains lacking either gene have much reduced m 7 G, and coexpression of both proteins is required to overproduce activity. Aniline cleavage mapping shows that Trm8/Trm82 proteins modify pre-tRNA Phe at G46, the site that is modified in vivo. Trm8 and Trm82 proteins form a complex, as affinity purification of Trm8 protein causes copurification of Trm82 protein in approximate equimolar yield. This functional two-protein family appears to be retained in eukaryotes, as expression of both corresponding human proteins, METTL1 and WDR4, is required for m 7 Gmethyltransferase activity.

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Molecular Analysis ofMETTL1,a Novel Human Methyltransferase-like Gene with a High Degree of Phylogenetic Conservation

Cited by 32 publications

References 8 publications

tRNA m⁷G methyltransferase Trm8p/Trm82p: Evidence linking activity to a growth phenotype and implicating Trm82p in maintaining levels of active Trm8p

tRNA m⁷G methyltransferase Trm8p/Trm82p: Evidence linking activity to a growth phenotype and implicating Trm82p in maintaining levels of active Trm8p

Transfer RNA Methytransferases and Their Corresponding Modifications in Budding Yeast and Humans: Activities, Predications, and Potential Roles in Human Health

Two proteins that form a complex are required for 7-methylguanosine modification of yeast tRNA

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Molecular Analysis ofMETTL1,a Novel Human Methyltransferase-like Gene with a High Degree of Phylogenetic Conservation

Cited by 32 publications

References 8 publications

tRNA m7G methyltransferase Trm8p/Trm82p: Evidence linking activity to a growth phenotype and implicating Trm82p in maintaining levels of active Trm8p

tRNA m7G methyltransferase Trm8p/Trm82p: Evidence linking activity to a growth phenotype and implicating Trm82p in maintaining levels of active Trm8p

Transfer RNA Methytransferases and Their Corresponding Modifications in Budding Yeast and Humans: Activities, Predications, and Potential Roles in Human Health

Two proteins that form a complex are required for 7-methylguanosine modification of yeast tRNA

Contact Info

Product

Resources

About

tRNA m⁷G methyltransferase Trm8p/Trm82p: Evidence linking activity to a growth phenotype and implicating Trm82p in maintaining levels of active Trm8p

tRNA m⁷G methyltransferase Trm8p/Trm82p: Evidence linking activity to a growth phenotype and implicating Trm82p in maintaining levels of active Trm8p