RNA polymerase III (Pol III) produces essential components of the biosynthetic machinery, and therefore its activity is tightly coupled with cell growth and metabolism. In the yeast Saccharomyces cerevisiae, Maf1 is the only known global and direct Pol III transcription repressor which mediates numerous stress signals. Here we demonstrate that transcription regulation by Maf1 is not limited to stress but is important for the switch between fermentation and respiration. Under respiratory conditions, Maf1 is activated by dephosphorylation and imported into the nucleus. The transition from a nonfermentable carbon source to that of glucose induces Maf1 phosphorylation and its relocation to the cytoplasm. The absence of Maf1-mediated control of tRNA synthesis impairs cell viability in nonfermentable carbon sources. The respiratory phenotype of maf1-⌬ allowed genetic suppression studies to dissect the mechanism of Maf1 action on the Pol III transcription apparatus. Moreover, in cells grown in a nonfermentable carbon source, Maf1 regulates the levels of different tRNAs to various extents. The differences in regulation may contribute to the physiological role of Maf1.
Maf1 protein is a global negative regulator of RNA polymerase (Pol) III transcription conserved from yeast to man. We report that phosphorylation of Maf1 by casein kinase II (CK2), a highly evolutionarily conserved eukaryotic kinase, is required for efficient Pol III transcription. Both recombinant human and yeast CK2 were able to phosphorylate purified human or yeast Maf1, indicating that Maf1 can be a direct substrate of CK2. Upon transfer of Saccharomyces cerevisiae from repressive to favorable growth conditions, CK2 activity is required for the release of Maf1 from Pol III bound to a tRNA gene and for subsequent activation of tRNA transcription. In a yeast strain lacking Maf1, CK2 inhibition showed no effect on tRNA synthesis, confirming that CK2 activates Pol III via Maf1. Additionally, CK2 was found to associate with tRNA genes, and this association is enhanced in absence of Maf1, especially under repressive conditions. These results corroborate the previously reported TFIIIB-CK2 interaction and indicate an important role of CK2-mediated Maf1 phosphorylation in triggering Pol III activation.RNA polymerase III regulation | transfer RNA | casein kinase II regulation R NA polymerase (Pol) III is responsible for the transcription of some 300 different genes in yeast (class III genes), mostly tRNA genes (1). In-depth analyses of the yeast Pol III transcription system have revealed a cascade of protein-DNA and protein-protein interactions leading to the recruitment of Pol III to its target tRNA genes: binding of the six-subunit TFIIIC factor to the intragenic promoter, TFIIIC-directed recruitment and assembly of the three subunits of TFIIIB (TBP, Brf1, and Bdp1) and subsequent recruitment of the 17-subunit Pol III enzyme (2). High rate of tRNA transcription is achieved through many rounds of reinitiation by Pol III on stable DNA-bound complexes of the initiation factor TFIIIB (3, 4).Pol III is under control of the general negative regulator Maf1 (5, 6), which binds to Pol III clamp and rearranges specific subcomplex C82/34/31, which is required for transcription initiation (7). In the repressive complex, Maf1 impairs recruitment of Pol III to a complex of promoter DNA with the initiation factors TFIIB and thus prevents closed-complex formation (4, 7). Maf1 is essential for repressing Pol III transcription in yeast and mediates several signaling pathways (8). In addition to the down-regulation that occurs normally in the stationary phase, Pol III repression accompanying starvation, respiratory growth, as well as oxidative and replication stress, also requires Maf1 (9-11). Maf1 inhibits Pol III transcription via a mechanism that depends on the dephosphorylation and nuclear accumulation of Maf1 followed by its physical association with Pol III at Pol III-transcribed genes genomewide (6, 12). In contrast Maf1 phosphorylation occurs in favorable growth conditions and is linked to cytoplasmic localization of Maf1 (6, 13).Maf1 was recently found to be phosphorylated by protein kinases PKA (14, 15), Sch9 (16-18), and TO...
Maf1 is negative regulator of RNA polymerase III in yeast. We observed high levels of both primary transcript and end-matured, intron-containing pre-tRNAs in the maf1⌬ strain. This pre-tRNA accumulation could be overcome by transcription inhibition, arguing against a direct role of Maf1 in tRNA maturation and suggesting saturation of processing machinery by the increased amounts of primary transcripts. Saturation of the tRNA exportin, Los1, is one reason why end-matured introncontaining pre-tRNAs accumulate in maf1⌬ cells. However, it is likely possible that other components of the processing pathway are also limiting when tRNA transcription is increased. According to our model, Maf1-mediated transcription control and nuclear export by Los1 are two major stages of tRNA biosynthesis that are regulated by environmental conditions in a coordinated manner.
The synthesis of transfer RNA (tRNA) is directed by RNA polymerase III (Pol III) specialized in high-level transcription of short DNA templates. Pol III recruitment to tRNA genes is controlled by two general initiation factors, TFIIIB and TFIIIC. They are multi-protein complexes regulated at the level of expression of individual subunits, as well as through phosphorylation and interaction with partner proteins. Here, we describe particular aspects of TFIIIB and TFIIIC control in yeast and human cells. Under stress conditions, tRNA synthesis is negatively regulated by the MAF1 protein, which interacts directly with Pol III. Sequence and function of MAF1 are conserved among eukaryotic organisms from yeast to humans. MAF1 is a phosphoprotein which mediates diverse regulatory signals to Pol III. Interestingly, there is a subset of housekeeping tRNA genes, both in the yeast and human genome, which are less sensitive to MAF1-dependent repression. The possible mechanisms responsible for this differential regulation of tRNA synthesis by MAF1 are discussed.
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