Recently, a role for the anticodon wobble uridine modification 5-methoxycarbonylmethyl-2-thiouridine (mcm⁵s²U) has been revealed in the suppression of translational +1 frameshifts in Saccharomyces cerevisiae. Loss of either the mcm⁵U or s²U parts of the modification elevated +1 frameshift rates and results obtained with reporters involving a tRNA^Lys_UUU dependent frameshift site suggested these effects are caused by reduced ribosomal A-site binding of the hypomodified tRNA. Combined loss of mcm⁵U and s²U leads to increased ribosome pausing at tRNA^Lys_UUU dependent codons and synergistic growth defects but effects on +1 frameshift rates remained undefined to this end. We show in here that simultaneous removal of mcm⁵U and s²U results in synergistically increased +1 frameshift rates that are suppressible by extra copies of tRNA^Lys_UUU. Thus, two distinct chemical modifications of the same wobble base independently contribute to reading frame maintenance, loss of which may cause or contribute to observed growth defects. Since the thiolation pathway is sensitive to moderately elevated temperatures in yeast, we observe a heat-induced increase of +1 frameshift rates in wild type cells that depends on the sulfur transfer protein Urm1. Furthermore, we find that temperature-induced frameshifting is kept in check by the dehydration of N6-threonylcarbamoyladenosine (t⁶A) to its cyclic derivative (ct⁶A) at the anticodon adjacent position 37. Since loss of ct⁶A in elp3 or urm1 mutant cells is detrimental for temperature stress resistance we assume that conversion of t⁶A to ct⁶A serves to limit deleterious effects on translational fidelity caused by hypomodified states of wobble uridine bases.

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Journal Article | Research Support, Non-U.S. Gov't

Authors

Klassen R, Bruch A, Schaffrath R

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