Should I stay or should I go: TFIIIC as assembly factor and barrier in RNA polymerase III transcription

Abstract

Critical for the regulation of eukaryotic gene transcription is the assembly and interplay of general transcription factors (GTFs) with RNA polymerases (RNAPs), leading to the formation of pre-initiation complexes (PICs) as a rate-limiting step in transcription activation. Compared with RNAPII PIC assembly involving many GTFs, activators, and co-activators, RNAPIII PIC assembly is less complex, involving mainly the four GTFs TFIIIA, TFIIIB, TFIIIC, and snRNA activating protein complex with only a few additional factors. The RNAPIII-specific GTF TFIIIC is present in type I and II promoters. One prominent area of investigation has been the dynamic interaction between TFIIIC and its promoter elements, the varying affinities of TFIIIC toward these elements, and the flexible linker within TFIIIC. Additionally, evidence suggests that TFIIIC may play a dual role, acting as an assembly factor that positions TFIIIB during PIC formation and as a barrier during RNAPIII-mediated transcription. By summarizing recent structural, biochemical, and genomic data, this review explores the mechanisms by which RNAPIII-specific GTFs, with a focus on TFIIIC, dynamically regulate RNAPIII transcription.

Keywords: RNA polymerase III; TFIIIC; general transcription factor; tRNA transcription; transcription initiation.

Figure 1. Available crystal and cryo-EM structures of individual RNAPIII GTFs and complexes bound to their promoter elements.

(A) Type I promoter. TFIIIA (ZF1–ZF6) bound to the C-box and IE (PDB: 1TF6); cryo-EM structure of TFIIIA-TFIIIC bound to the 5S RNA gene, where PDB:8FFZ was fitted into the corresponding density map (EMD-29358) with unsupported regions removed; cryo-EM structure of TFIIIA-TFIIIC-Brf1-TBP bound to the 5S RNA gene, which wraps around the complex (PDB:8FFZ). (B) Type II promoter. Left panel: Cryo-EM structures of human TFIIIC bound to a tRNA gene include the unbound τA subcomplex (PDB:8CLK) and the τB subcomplex bound to the B-box (PDB:8CLI). Right panel: Fully engaged yeast TFIIIC bound to a tRNA gene is represented by cryo-EM structures of the τA subcomplex (PDB:9GCK) and the τB subcomplex (PDB:9GC3). So far, no structural information about a TFIIIC-TFIIIB complex has been reported. The complete yeast TFIIIC–DNA model was built by positioning a 45-bp tRNA^His DNA duplex into the τA–DNA map and a 40-bp segment into the τB–DNA map, with orientations and distances refined using cryo-EM single-particle mapping described in [5]. (C) Type III promoter. Cryo-EM structure of miniSNAPc bound to the PSE motif (PDB: 7XUR); crystal structure of TFIIIB (Brf2 type) bound to its TATA box site (PDB: 5N9G).

Figure 2. The dynamics of TFIIIC during RNAPIII transcription initiation.

(1) The TFIIIC submodule τB binds with high affinity to the B-box and anchors to the tRNA gene. (2) This facilitates subsequent recruitment of τA to the A-box with weaker affinity. To bridge the region between the A- and B-box, TFIIIC can simultaneously engage with both promoter elements due to a flexible linker in subunit TFIIIC220 in human and subunit τ138 in yeast for spacings <74 bp. Subunit TFIIIC220 in human TFIIIC and yeast τ138 take part in both the τA and the τB subcomplexes, with its N-terminus belonging to τB and its C-terminus to τA. When the intervening sequence between the A- and B-box exceeds 74 bp, TFIIIC could adapt by inducing DNA looping. (3) Once TFIIIC is assembled on the tRNA gene, TFIIIB is recruited and placed upstream the TSS. (4) Whether TFIIIC is subsequently fully released from the tRNA gene remains unclear. Two hypotheses have been proposed: 1) The τA lobe gets detached upon interaction with TFIIIB or 2) interaction with RNAPIII disrupts the interaction between τA and τB to promote TFIIIC dissociation. TSS, transcription start site; TES, transcription end site.

Figure 3. A two-state model for TFIIIC.

In the active state (left), when RNAPIII elongation takes place, at least τA dissociates from the tRNA gene, which allows RNAPIII to move along the tRNA gene to perform transcription elongation. Possibly, τB still interacts with the tRNA gene to interfere with elongating RNAPIII by acting as a roadblock. Upon repressive signals (right), RNAPIII occupancy at the tRNA is decreased and is bound by Maf1. In addition, TFIIIC occupancy on the chromatin is increased. The interaction between TFIIIC and TFIIIB is increased, and possibly TFIIIB remains in close proximity to the tRNA gene in an inactive state to quickly resume transcription upon changing to more favorable conditions. TFIIIC could act as a transient barrier or as an active repressor (or activator) by recruitment of repressive (or active) marks.