Yeast Killer Toxins: Fundamentals and Applications

Meinhardt, Friedhelm; Klassen, Roland

doi:10.1007/978-3-642-00286-1_6

Cited by 19 publications

(21 citation statements)

References 216 publications

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“…At the same time, Kre1p was shown to be necessary for the action of the K1 toxin on target cells and was regarded as a plasma membrane-localized receptor. However, the discovery of numerous yeast factors involved in K1 resistance and their connection to cell wall and plasma membrane biogenesis suggests that other targets of K1 cannot be excluded (9, 11).While dominant in vineyard/winery ecosystems, K2 toxin was studied less extensively (5,(12)(13)(14). It was generally assumed that K1 and K2 toxins act in a similar fashion (11,15,16).…”

mentioning

confidence: 99%

“…While dominant in vineyard/winery ecosystems, K2 toxin was studied less extensively (5,(12)(13)(14). It was generally assumed that K1 and K2 toxins act in a similar fashion (11,15,16).…”

mentioning

confidence: 99%

“…The toxins have been grouped into four types (K1, K2, K28, and Klus) based on their killing profiles and lack of crossimmunity (3,4). Such proteins are able to kill the nonkiller yeast, as well as yeast of other killer types, while the toxin-producing cells remain immune to their own or to the same type of killers (4,5). K1 toxin disrupts the regulated ion flux across the plasma membrane, leading to the death of sensitive yeast strains (6,7).…”

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confidence: 99%

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Yeast β-1,6-Glucan Is a Primary Target for the Saccharomyces cerevisiae K2 Toxin

et al. 2015

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b Certain Saccharomyces cerevisiae strains secrete different killer proteins of double-stranded-RNA origin. These proteins confer a growth advantage to their host by increasing its survival. K2 toxin affects the target cell by binding to the cell surface, disrupting the plasma membrane integrity, and inducing ion leakage. In this study, we determined that K2 toxin saturates the yeast cell surface receptors in 10 min. The apparent amount of K2 toxin, bound to a single cell of wild type yeast under saturating conditions, was estimated to be 435 to 460 molecules. It was found that an increased level of ␤-1,6-glucan directly correlates with the number of toxin molecules bound, thereby impacting the morphology and determining the fate of the yeast cell. We observed that the binding of K2 toxin to the yeast surface receptors proceeds in a similar manner as in case of the related K1 killer protein. It was demonstrated that the externally supplied pustulan, a poly-␤-1,6-glucan, but not the glucans bearing other linkage types (such as laminarin, chitin, and pullulan) efficiently inhibits the K2 toxin killing activity. In addition, the analysis of toxin binding to the intact cells and spheroplasts confirmed that majority of K2 protein molecules attach to the ␤-1,6-glucan, rather than the plasma membrane-localized receptors. Taken together, our results reveal that ␤-1,6-glucan is a primary target of K2 toxin and is important for the execution of its killing property.T he production of antimycotic killer toxins has been observed in several yeast genera and proved to be a widespread phenomenon (1, 2). Killer strains of Saccharomyces cerevisiae secrete protein toxins derived from a family of double-stranded RNAs (dsRNAs). The toxins have been grouped into four types (K1, K2, K28, and Klus) based on their killing profiles and lack of crossimmunity (3, 4). Such proteins are able to kill the nonkiller yeast, as well as yeast of other killer types, while the toxin-producing cells remain immune to their own or to the same type of killers (4, 5). K1 toxin disrupts the regulated ion flux across the plasma membrane, leading to the death of sensitive yeast strains (6, 7). The killing action of K1 toxin involves at least two steps. During the first step, the toxin binds to the cell wall, whereas the second step leads to the translocation and insertion of the toxin into the plasma membrane (6). Beta-1,6-glucan was originally proposed to be a cell wall receptor for K1 (8). Analysis of several kre mutants demonstrated that decrease of the cell wall ␤-1,6-glucan level leads to K1 resistance, thus confirming the involvement of this type of glucan in toxin binding (9). During the second step, K1 toxin interacts with plasma membrane receptors and disrupts the functional integrity of the plasma membrane either by inducing the formation of new ion channels (7) or through the activation of existing potassium channels (10). Products of TOK1 (protein, forming the potassium ion channel) and KRE1 (glycoprotein, involved in ␤-glucan assembly) have be...

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confidence: 99%

“…While dominant in vineyard/winery ecosystems, K2 toxin was studied less extensively (5,(12)(13)(14). It was generally assumed that K1 and K2 toxins act in a similar fashion (11,15,16).…”

mentioning

confidence: 99%

mentioning

confidence: 99%

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Yeast β-1,6-Glucan Is a Primary Target for the Saccharomyces cerevisiae K2 Toxin

et al. 2015

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“…A large variety of yeast secrete different classes of compounds aimed at preventing the growth of the competitors (Meinhardt, Klassen, 2009). These compounds include small molecules, antibiotic peptides, and even proteins.…”

Section: Introductionmentioning

confidence: 99%

A quick and reliable method for genome-wide host factor screening of Saccharomyces cerevisiae killer toxins

Servienė¹,

Lukša²,

Vepštaitė-Monstavičė³

et al. 2017

biologija

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Numerous yeasts produce toxic compounds to fight the competitors. Such compounds include small molecules (like antibiotics), antibiotic peptides, and also larger proteins, including killer toxins. Their ability to affect the cell depends on the host factors modulating the killing activity. Here we describe a robotics-based method to advance the genome-wide screening for the host factors affecting sensitivity of budding yeast to the killer toxins using the K2 system as the model. We demonstrate that arraying the mutant library on the agar plates containing the K2 killer toxinproducing strain and/or purified toxin ("survival" assay) increases the sensitivity and speed of the screen and decreases the costs compared to the traditional "killer" assay. We show the applicability of a new screening method of searching for the host factors using a killer strain isolated from agricultural plant environment. In addition, the "survival" assay allows identification of previously undetected factors that could be the "missing links" in the pathways of toxininduced cellular responses.

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“…Zymocin and PaT arrest growth of the nonself yeast species Saccharomyces cerevisiae by exposure from without. The a and b subunits of zymocin and the PaOrf1 subunit of PaT interact with the target cell surface to effect the transport of the g subunit and the PaOrf2 subunit (the anticodon endonuclease enzymes) into the cytoplasm of the target cell (Jablonowski et al 2004;Jablonowski and Schaffrath 2007;Meinhardt and Klassen 2009). Toxicity can also be achieved from within by induced intracellular expression in S. cerevisiae of just the g-toxin or PaOrf2 subunits (minus their N-terminal signal peptides) (Butler et al 1991;Klassen et al 2004).…”

Section: Introductionmentioning

confidence: 99%

A fungal anticodon nuclease ribotoxin exploits a secondary cleavage site to evade tRNA repair

Meineke

Alene

Schwer

et al. 2012

RNA

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PaOrf2 and g-toxin subunits of Pichia acaciae toxin (PaT) and Kluyveromyces lactis zymocin are tRNA anticodon nucleases. These secreted ribotoxins are assimilated by Saccharomyces cerevisiae, wherein they arrest growth by depleting specific tRNAs. Toxicity can be recapitulated by induced intracellular expression of PaOrf2 or g-toxin in S. cerevisiae. Mutational analysis of g-toxin has identified amino acids required for ribotoxicity in vivo and RNA transesterification in vitro. Here, we report that PaOrf2 residues Glu9 and His287 (putative counterparts of g-toxin Glu9 and His209) are essential for toxicity. Our results suggest a similar basis for RNA transesterification by PaOrf2 and g-toxin, despite their dissimilar primary structures and distinctive tRNA target specificities. PaOrf2 makes two sequential incisions in tRNA, the first of which occurs 39 from the mcm 5 s 2 U wobble nucleoside and depends on mcm 5 . A second incision two nucleotides upstream results in the net excision of a di-nucleotide. Expression of phage and plant tRNA repair systems can relieve PaOrf2 toxicity when tRNA cleavage is restricted to the secondary site in elp3 cells that lack the mcm 5 wobble U modification. Whereas the endogenous yeast tRNA ligase Trl1 can heal tRNA halves produced by PaOrf2 cleavage in elp3 cells, its RNA sealing activity is inadequate to complete the repair. Compatible sealing activity can be provided in trans by plant tRNA ligase. The damage-rescuing ability of tRNA repair systems is lost when PaOrf2 can break tRNA at both sites. These results highlight the logic of a two-incision mechanism of tRNA anticodon damage that evades productive repair by tRNA ligases.

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Yeast Killer Toxins: Fundamentals and Applications

Cited by 19 publications

References 216 publications

Yeast β-1,6-Glucan Is a Primary Target for the Saccharomyces cerevisiae K2 Toxin

Yeast β-1,6-Glucan Is a Primary Target for the Saccharomyces cerevisiae K2 Toxin

A quick and reliable method for genome-wide host factor screening of Saccharomyces cerevisiae killer toxins

A fungal anticodon nuclease ribotoxin exploits a secondary cleavage site to evade tRNA repair

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