The primary target of the killer toxin from <i>Pichia acaciae</i> is tRNA<sup>Gln</sup>

Klassen, Roland; Paluszynski, John P.; Wemhoff, Sabrina; Pfeiffer, Annika; Fricke, Julia; Meinhardt, Friedhelm

doi:10.1111/j.1365-2958.2008.06319.x

Cited by 67 publications

(118 citation statements)

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“…Consequent depletion of the tRNA Gln(UUG) pool arrests yeast growth (18). Galactose-induced expression in S. cerevisiae of an intracellular form of PaT recapitulates its toxicity (18). The salient finding here was that replacing yeast Trl1 with RtcB as the source of tRNA ligase protected S. cerevisiae from PaT-mediated growth inhibition (Fig.…”

Section: Resultsmentioning

confidence: 70%

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RtcB, a Novel RNA Ligase, Can Catalyze tRNA Splicing and HAC1 mRNA Splicing in Vivo

Tanaka

Meineke

Shuman

2011

Journal of Biological Chemistry

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RtcB enzymes are novel RNA ligases that join 2,3-cyclic phosphate and 5-OH ends. The phylogenetic distribution of RtcB points to its candidacy as a tRNA splicing/repair enzyme. Here we show that Escherichia coli RtcB is competent and sufficient for tRNA splicing in vivo by virtue of its ability to complement growth of yeast cells that lack the endogenous "healing/ sealing-type" tRNA ligase Trl1. RtcB also protects yeast trl1⌬ cells against a fungal ribotoxin that incises the anticodon loop of cellular tRNAs. Moreover, RtcB can replace Trl1 as the catalyst of HAC1 mRNA splicing during the unfolded protein response. Thus, RtcB is a bona fide RNA repair enzyme with broad physiological actions. Biochemical analysis of RtcB highlights the uniqueness of its active site and catalytic mechanism. Our findings draw attention to tRNA ligase as a promising drug target.Escherichia coli RtcB exemplifies a new family of RNA ligases that directly seal 2Ј,3Ј-cyclic phosphate and 5Ј-OH ends (1-3). Direct ligation is thought to be the main pathway of tRNA splicing in animals and archaea (4 -6). By contrast, yeast and plants rely on a different mechanism of tRNA splicing in which the broken 3Ј and 5Ј ends are healed (converted to a 3Ј-OH/2Ј-PO 4 and 5Ј-PO 4 , respectively) and then sealed by a classical ATPdependent RNA ligase (7) (see Fig. 1). RNA ligases of the RtcB family are present in metazoa and protozoa, but not in fungi and plants. RtcB homologs purified from archaeal and mammalian cells can seal broken tRNA halves and are thereby imputed to be the catalysts of archaeal and mammalian tRNA splicing (2, 3). However, this scenario is complicated by the existence of a yeast/plant-like tRNA splicing pathway in mammalian cells (8 -12) and of analogous yeast-like RNA repair enzymes in many archaeal taxa (13-16). Definitive genetic evidence that RtcB is the sole essential agent of the repair phase of mammalian or archaeal tRNA splicing is lacking, and the available genetic evidence concerning the healing-sealing pathway in animals is equivocal. Genetic ablation of a murine homolog of a yeast-like pathway component Tpt1 (the enzyme that removes the 2Ј-phosphate at the splice junction; see Fig. 1) has no discernible phenotype (17), suggesting that the mammalian yeastlike pathway either is functionally redundant with direct ligation or is non-contributory to mammalian tRNA splicing. By contrast, siRNA-directed depletion of the mammalian RNA 5Ј-kinase (an ortholog of the kinase domain of yeast/plant tRNA ligase) elicited a defect in tRNA splicing in vitro (10). However, siRNA-directed depletion of mammalian RtcB was also reported to inhibit ligation of tRNA halves in cell extracts and to delay tRNA splicing in living cells (3). These findings leave unresolved the following key issues: (i) whether RtcB can suffice for tRNA splicing as the only source of tRNA ligase activity in a eukaryal cell and (ii) whether RtcB can perform other RNA repair functions in vivo. Here we address these questions by using budding yeast as a surrogate geneti...

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Section: Resultsmentioning

confidence: 70%

“…PaT is a secreted fungal defense molecule that penetrates S. cerevisiae cells and, upon accessing the cytoplasm, breaks the anticodon loop of tRNA Gln(UUG) . Consequent depletion of the tRNA Gln(UUG) pool arrests yeast growth (18). Galactose-induced expression in S. cerevisiae of an intracellular form of PaT recapitulates its toxicity (18).…”

Section: Resultsmentioning

confidence: 99%

RtcB, a Novel RNA Ligase, Can Catalyze tRNA Splicing and HAC1 mRNA Splicing in Vivo

Tanaka

Meineke

Shuman

2011

Journal of Biological Chemistry

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“…Killer toxins from the genus Pichia have heterogeneous mechanisms of activity; some of them have similar characteristics to that determined for the P. membranifaciens PMKT (Santos & Marquina, 2004a;Pfeiffer & Radler, 1984;Middelbeek et al, 1979), and others do not. PaT, the killer toxin produced by P. acaciae, has a tRNase activity (Klassen et al, 2008) whereas P. anomala NCYC 434 has an exo-b-1,3-glucanase activity against sensitive yeast strains (Izgü et al, 2007).…”

Section: Introductionmentioning

confidence: 99%

“…Strains of Pichia membranifaciens are common contaminants in food-related environments and occur with high frequency in fermenting olive brines (Marquina et al, 1992(Marquina et al, , 1997. Within the genus Pichia, which is heterogeneous from a taxonomic point of view (Kurtzman & Fell, 1998;Kurtzman & Robnett, 1998), P. acaciae (McCracken et al, 1994;Klassen et al, 2008), P. anomala (Comitini et al, 2004;Wang, et al, 2007;Izgü et al, 2007), P. farinosa (Suzuki & Nikkuni, 1994), P. inositovora (Klassen & Meinhardt, 2003) and P. kluyveri (Middelbeek et al, 1979) produce different killer toxins. Killer toxins from the genus Pichia have heterogeneous mechanisms of activity; some of them have similar characteristics to that determined for the P. membranifaciens PMKT (Santos & Marquina, 2004a;Pfeiffer & Radler, 1984;Middelbeek et al, 1979), and others do not.…”

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confidence: 99%

PMKT2, a new killer toxin from Pichia membranifaciens, and its promising biotechnological properties for control of the spoilage yeast Brettanomyces bruxellensis

et al. 2009

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Pichia membranifaciens CYC 1086 secretes a killer toxin (PMKT2) that is inhibitory to a variety of spoilage yeasts and fungi of agronomical interest. The killer toxin in the culture supernatant was concentrated by ultrafiltration and purified to homogeneity by two successive steps, including native electrophoresis and HPLC gel filtration. Biochemical characterization of the toxin showed it to be a protein with an apparent molecular mass of 30 kDa and an isoelectric point of 3.7. At pH 4.5, optimal killer activity was observed at temperatures up to 20 6C. Above approximately this pH, activity decreased sharply and was barely noticeable at pH 6. The toxin concentrations present in the supernatant during optimal production conditions exerted a fungicidal effect on a variety of fungal and yeast strains. The results obtained suggest that PMKT2 has different physico-chemical properties from PMKT as well as different potential uses in the biocontrol of spoilage yeasts. PMKT2 was able to inhibit Brettanomyces bruxellensis while Saccharomyces cerevisiae was fully resistant, indicating that PMKT2 could be used in wine fermentations to avoid the development of the spoilage yeast without deleterious effects on the fermentative strain. In small-scale fermentations, PMKT2, as well as P. membranifaciens CYC 1086, was able to inhibit B. bruxellensis, verifying the biocontrol activity of PMKT2 in simulated winemaking conditions. INTRODUCTIONWorldwide, microbial growth destroys large amounts of various products, causing yield losses in the agronomical and biotechnological industries. Traditionally, biocides have been used to deal with these problems but different disadvantages such as establishment of resistant strains and suppression of natural competitors have made alternatives such as biological control necessary (Beever et al., 1989;Raposo et al., 2000). Biological control strategies include natural plant-and animal-derived compounds, as well as antagonistic micro-organisms (Ciani & Fatichenti, 2001).During recent decades, microbiological control of spoilage micro-organisms has evolved as a possibility. Many yeast strains and other micro-organisms inhibiting plant pathogens have been reported, especially within the fruit-and vegetable-producing sector, and several new products have reached the commercial market (Janisiewicz & Korsten, 2002). The suggested modes of action of biocontrol yeasts are not likely to constitute any hazard for the consumer (Janisiewicz et al., 2001;Masih & Paul, 2002;Comitini et al., 2004;Santos & Marquina, 2004a).The food and beverage industries were among the first to explore the application of killer-toxin-producing yeasts to kill spoilage micro-organisms (Lowes et al., 2000). Yeast strains often achieve competitive advantage by producing killer toxins, which kill off competing sensitive cells belonging to either the same or a different species (Young, 1987;Ciani & Fatichenti, 2001). The most thoroughly studied examples are the Saccharomyces cerevisiae toxins K1, K2 and K28; producers of these toxi...

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“…This process is also influenced by modifications -yeast Trm9p, and mammalian NSUN2 and DNMT2 have all been implicated in regulating cleavage through their methyltransferase activity. 32,[63][64][65] The resulting fragments have been demonstrated to bind and inhibit translation machinery, 66 as well as bind and destabilize mRNAs directly. 67 In mouse epidermal stem cells, the expression level of the RNA m 5 C methyltransferase NSUN2 is low but increases over the course of differentiation.…”

Section: Trnamentioning

confidence: 99%

Publisher's Note

2017

RNA Biology

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RNA modifications have long been known to be central in the proper function of tRNA and rRNA. While chemical modifications in mRNA were discovered decades ago, their function has remained largely mysterious until recently. Using enrichment strategies coupled to next generation sequencing, multiple modifications have now been mapped on a transcriptome-wide scale in a variety of contexts. We now know that RNA modifications influence cell biology by many different mechanisms -by influencing RNA structure, by tuning interactions within the ribosome, and by recruiting specific binding proteins that intersect with other signaling pathways. They are also dynamic, changing in distribution or level in response to stresses such as heat shock and nutrient deprivation. Here, we provide an overview of recent themes that have emerged from the substantial progress that has been made in our understanding of chemical modifications across many major RNA classes in eukaryotes.

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The primary target of the killer toxin from Pichia acaciae is tRNA^Gln

Cited by 67 publications

References 43 publications

RtcB, a Novel RNA Ligase, Can Catalyze tRNA Splicing and HAC1 mRNA Splicing in Vivo

RtcB, a Novel RNA Ligase, Can Catalyze tRNA Splicing and HAC1 mRNA Splicing in Vivo

PMKT2, a new killer toxin from Pichia membranifaciens, and its promising biotechnological properties for control of the spoilage yeast Brettanomyces bruxellensis

Publisher's Note

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The primary target of the killer toxin from Pichia acaciae is tRNAGln

Cited by 67 publications

References 43 publications

RtcB, a Novel RNA Ligase, Can Catalyze tRNA Splicing and HAC1 mRNA Splicing in Vivo

RtcB, a Novel RNA Ligase, Can Catalyze tRNA Splicing and HAC1 mRNA Splicing in Vivo

PMKT2, a new killer toxin from Pichia membranifaciens, and its promising biotechnological properties for control of the spoilage yeast Brettanomyces bruxellensis

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The primary target of the killer toxin from Pichia acaciae is tRNA^Gln