Casein kinase II-mediated phosphorylation of general repressor Maf1 triggers RNA polymerase III activation

Abstract

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...

Recovery of RNA Polymerase III Transcription from the Glycerol-repressed State

“…Although the details of its involvement in this process are not fully understood, CK2 interacts with and phosphorylates subunits of the initiation factor TFIIIB (TBP in yeast and Brf1 in humans) and stimulates the assembly of TFIIIB on pol III gene promoters (19,20). CK2 is also associated with pol III genes in ChIP experiments (21,24,25), and therefore has the potential to regulate ongoing transcription from complexes assembled on the DNA. Previous work in human cells indicates that this is important during mitotic repression of pol III transcription where CK2 phosphorylation of the Bdp1 subunit of TFIIIB leads to its dissociation from chromatin (24).…”

Maf1, repressor of tRNA transcription, is involved in the control of gluconeogenetic genes in Saccharomyces cerevisiae

“…Yeast strain YPH500 (MATα ura3-52 lys2-801_amber ade2-101_ochre trp1-Δ63 his3-Δ200 leu2-Δ) and its isogenic derivatives carrying maf1::KanMX4 or maf1::URA3 (maf1Δ hereafter) were described previously (Boguta et al, 1997;Graczyk et al, 2011).…”

“…Maf1 is regulated by its phosphorylation-dependent cellular localization (Boisnard et al, 2009;Graczyk et al, 2011;Huber et al, 2009;Lee et al, 2009;Moir et al, 2006; A C C E P T E D M A N U S C R I P T…”

Does casein kinase II phosphorylation of Maf1 trigger RNA polymerase III activation?

“…The recent study of Graczyk et al (1) reported a role for casein kinase II (CK2) phosphorylation of Maf1 during the reactivation of RNA polymerase III (Pol III) transcription that occurs as cells transition from repressive to favorable growth conditions. This is a potentially important finding.…”

“…Using the differential mobility of Maf1 in one-dimensional SDS gels, Graczyk et al (1) reported changes in Maf1 phosphorylation upon conditional or pharmacological inhibition of CK2. However, prior work has established that these same changes in the mobility of Maf1 can be caused by mutation of as few as two phosphorylation sites (serines 177 and 178), sites targeted by protein kinase A and Sch9 that are not part of a CK2 recognition motif (2).…”