Why Are tRNAs Overproduced in the Absence of Maf1, a Negative Regulator of RNAP III, Not Fully Functional?

tRNA biosynthesis in the eukaryotic cell is a multistep pathway, involving transcription, 5' and 3' end maturation, intron removal, and numerous modifications of nucleotides. Most of the genes coding for each of the elements essential for tRNA biosynthetic activities were primarily identified by genetic selection in yeast [1]. In these studies, the parental strain contained a tRNA gene that had been converted to a nonsense suppressor. Given the appropriate genetic background, phenotypic loss of suppression was used to select mutants producing non-functional tRNA. Among other proteins controlling tRNA biosynthesis, this approach led to identification of Maf1, a global repressor of tRNA transcription that is activated in response to stress. Themaf1-1mutant was originally selected in a genetic screen for decreased efficiency of tRNA suppressor SUP11 (tRNA Tyr/UAA) in budding yeast, Saccharomyces cerevisiae [2]. The role of Maf1 has been suggested by tRNA accumulation inmaf1Δ cells, observed regardless of the repressive growth conditions [3]. An analogous decrease of tRNA-suppressor (tRNA Ser/ UCA) activity was detected for themaf1Δmutant in Schizosaccharomyces pombe [4]. Interestingly, the effect of Maf1 on the efficiency of tRNA-mediated suppression is contrary to that expected. Although one would assume that increased cellular tRNA levels should improve the efficiency of tRNA-mediated nonsense suppression, data show the opposite is true. Despite nearly two decades since the original discovery, the mechanism by which tRNA accumulation in themaf1Δmutant leads to the antisuppressor phenotype is still not understood. The simplest hypothesis is that tRNAs transcribed inmaf1Δ cells are incompletely processed, hypomodified, or fail to be appropriately delivered to ribosomes. It is worth noting that both primary transcripts and end-processed, intron-containing tRNA precursors were abnormally abundant in the absence of Maf1, and the nuclear export machinery was overloaded [5]. It was, however, unknown which processes in the tRNA maturation pathway were saturated by the increased amounts of primary transcripts in cells lacking Maf1. The current study by Arimbasseri and colleagues [4] solves a long-term conundrum: why tRNAs overproduced in the absence of Maf1 are not fully functional. Their elegant work makes a convincing case for the saturation of the dimethyltransferase Trm1 playing a crucial role in the mechanism by which Maf1 affects tRNA suppression. By using tRNA-HydroSeq technique to examine tRNA modification levels in S. pombe on the global scale, the authors have shown that Trm1 substrates are not fully modified even in wild type cells. Further decrease of Trm1-mediated G26 dimethylation on certain tRNAs was detected in themaf1Δmutant. Consequently, hypomodification of G26 due to limited Trm1 reduces the activity of tRNA-suppressor-Ser/UCA and accounts for antisuppression. This hypothesis was validated by genetic complementation of the antisuppressor phenotype of themaf1Δmutant in fission yeast by