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The primary target of the killer toxin from Pichia acaciae is tRNA Gln
Author(s) -
Klassen Roland,
Paluszynski John P.,
Wemhoff Sabrina,
Pfeiffer Annika,
Fricke Julia,
Meinhardt Friedhelm
Publication year - 2008
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/j.1365-2958.2008.06319.x
Subject(s) - biology , transfer rna , protein subunit , microbiology and biotechnology , biochemistry , rna , gene
Summary The Pichia acaciae killer toxin (PaT) arrests yeast cells in the S‐phase of the cell cycle and induces DNA double‐strand breaks (DSBs). Surprisingly, loss of the tRNA‐methyltransferase Trm9 – along with the Elongator complex involved in synthesis of 5‐methoxy‐carbonyl‐methyl (mcm 5 ) modification in certain tRNAs – conferred resistance against PaT. Overexpression of mcm 5 ‐modified tRNAs identified tRNA Gln (UUG) as the intracellular target. Consistently, toxin‐challenged cells displayed reduced levels of tRNA Gln and in vitro the heterologously expressed active toxin subunit disrupts the integrity of tRNA Gln (UUG) . Other than Kluyveromyces lactis zymocin, an endonuclease specific for tRNA Glu (UUC) , affecting its target in a mcm 5 ‐dependent manner, PaT exerts activity also on tRNA Gln lacking such modification. As sensitivity is restored in trm9 elp3 double mutants, target tRNA cleavage is selectively inhibited by incomplete wobble uridine modification, as seen in trm9 , but not in elp3 or trm9 elp3 cells. In addition to tRNA Gln (UUG) , tRNA Gln (CUG) is also cleaved in vitro and overexpression of the corresponding gene increased resistance. Consistent with tRNA Gln (CUG) as an additional TRM9 ‐independent target, overexpression of PaT's tRNase subunit abolishes trm9 resistance. Most interestingly, a functional DSB repair pathway confers PaT but also zymocin resistance, suggesting DNA damage to occur generally concomitant with specific tRNA offence.