Maf1 is a conserved inhibitor of RNA polymerase III (Pol III) that influences phenotypes ranging from metabolic efficiency to lifespan. Here, we present a 3.3-Å-resolution cryo-EM structure of yeast Maf1 bound to Pol III, establishing that Maf1 sequesters Pol III elements involved in transcription initiation and binds the mobile C34 winged helix 2 domain, sealing off the active site. The Maf1 binding site overlaps with that of TFIIIB in the preinitiation complex.
Subscribe to Journal
Get full journal access for 1 year
only $18.75 per issue
All prices are NET prices.
VAT will be added later in the checkout.
Rent or Buy article
Get time limited or full article access on ReadCube.
All prices are NET prices.
The cryo-EM map of the Maf1–Pol III complex has been deposited to the Electron Microscopy Data Bank (EMDB) under the accession code EMD-10595. The coordinates of the corresponding model have been deposited to the Protein Data Bank (PDB) under accession code 6TUT. Source data for Fig. 2b,c are available with the paper.
Moir, R. D. & Willis, I. M. Regulation of pol III transcription by nutrient and stress signaling pathways. Biochim. Biophys. Acta Gene Regul. Mech. 1829, 361–375 (2013).
Grewal, S. S. Why should cancer biologists care about tRNAs? tRNA synthesis, mRNA translation and the control of growth. Biochim. Biophys. Acta 1849, 898–907 (2015).
Zhong, Q. et al. The significance of Brf1 overexpression in human hepatocellular carcinoma. Oncotarget 7, 6243–6254 (2016).
Gouge, J. et al. Redox signaling by the RNA polymerase III TFIIB-related factor Brf2. Cell 163, 1375–1387 (2015).
Palian, B. M. et al. Maf1 is a novel target of PTEN and PI3K signaling that negatively regulates oncogenesis and lipid metabolism. PLoS Genet. 10, e1004789 (2014).
Willis, I. M. & Moir, R. D. Signaling to and from the RNA polymerase III transcription and processing machinery. Annu. Rev. Biochem. 87, 75–94 (2018).
Willis, I. M. Maf1 phenotypes and cell physiology. Biochim. Biophys. Acta Gene Regul. Mech. 1861, 330–337 (2018).
Boguta, M. & Leniewska, E. Novel layers of RNA polymerase III control affecting tRNA gene transcription in eukaryotes. Open Biol 7, 170001 (2017).
Shetty, M. Maf1-dependent transcriptional regulation of tRNAs prevents genomic instability and is associated with extended lifespan. Aging Cell https://doi.org/10.1111/acel.13068 (2019).
Cai, Y. & Wei, Y. Stress resistance and lifespan are increased in C. elegans but decreased in S. cerevisiae by mafr-1/maf1 deletion. Oncotarget 7, 10812–10826 (2016).
Filer, D. et al. RNA polymerase III limits longevity downstream of TORC1. Nature 552, 263–267 (2017).
Bonhoure, N. et al. Loss of the RNA polymerase III repressor MAF1 confers obesity resistance. Genes Dev. 29, 934–947 (2015).
Willis, I. M., Moir, R. D. & Hernandez, N. Metabolic programming a lean phenotype by deregulation of RNA polymerase III. Proc. Natl Acad. Sci. USA 115, 12182–12187 (2018).
Bonhoure, N. et al. Chronic repression by MAF1 supports futile RNA cycling as a mechanism for obesity resistance. Preprint at bioRxiv https://doi.org/10.1101/775353 (2019).
Chen, C. Y. et al. Maf1 and repression of RNA polymerase III-mediated transcription drive adipocyte differentiation. Cell Rep. 24, 1852–1864 (2018).
Vannini, A. et al. Molecular basis of RNA polymerase III transcription repression by Maf1. Cell 143, 59–70 (2010).
Soprano, A. S. et al. Crystal structure and regulation of the citrus Pol III repressor MAF1 by auxin and phosphorylation. Structure 25, 1360–1370.e4 (2017).
Moir, R. D., Lee, J. & Willis, I. M. Recovery of RNA polymerase III transcription from the glycerol-repressed state. J. Biol. Chem. 287, 30833–30841 (2012).
Abascal-Palacios, G., Ramsay, E. P., Beuron, F., Morris, E. & Vannini, A. Structural basis of RNA polymerase III transcription initiation. Nature 553, 301–306 (2018).
Vorländer, M. K., Khatter, H., Wetzel, R., Hagen, W. J. H. & Müller, C. W. Molecular mechanism of promoter opening by RNA polymerase III. Nature 553, 295–300 (2018).
Han, Y., Yan, C., Fishbain, S., Ivanov, I. & He, Y. Structural visualization of RNA polymerase III transcription machineries. Cell Discov. 4, 40 (2018).
Pluta, K. et al. Maf1p, a negative effector of RNA polymerase III in Saccharomyces cerevisiae. Mol. Cell. Biol. 21, 5031–5040 (2001).
Moreno-Morcillo, M. et al. Solving the RNA polymerase I structural puzzle. Acta Crystallogr. D Biol. Crystallogr. 70, 2570–2582 (2014).
Male, G. et al. Architecture of TFIIIC and its role in RNA polymerase III pre-initiation complex assembly. Nat. Commun. 6, 7387 (2015).
Tegunov, D. & Cramer, P. Real-time cryo-electron microscopy data preprocessing with Warp. Nat. Methods 16, 1146–1152 (2019).
Kelly, L. A., Mezulis, S., Yates, C., Wass, M. & Sternberg, M. The Phyre2 web portal for protein modelling, prediction, and analysis. Nat. Protoc. 10, 845–858 (2015).
Brown, A. et al. Tools for macromolecular model building and refinement into electron cryo-microscopy reconstructions. Acta Crystallogr. D Biol. Crystallogr. 71, 136–153 (2015).
Adams, P. D. et al. PHENIX: a comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr. D Biol. Crystallogr. 66, 213–221 (2010).
Liao, Y., Moir, R. D. & Willis, I. M. Interactions of Brf1 peptides with the tetratricopeptide repeat-containing subunit of TFIIIC inhibit and promote preinitiation complex assembly. Mol. Cell. Biol. 26, 5946–5956 (2006).
This work was supported by an ERC Advanced Grant (ERC-2013-AdG340964-POL1PIC) to C.W.M. and R.W., a National Institutes of Health Grant (GM120358) to I.M.W. and an EMBL International PhD program award to M.K.V. We thank M. Girbig for help with transcription assays and critical reading of the manuscript and F. Weis for EM support. We are grateful to T. Hoffmann and J. Pecar for maintaining the high performance computing environment for EM data processing at EMBL.
The authors declare no competing interests.
Peer review information Beth Moorefield and Inês Chen were the primary editors on this article and managed its editorial process and peer review in collaboration with the rest of the editorial team.
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Representative micrograph denoised using Warp25, 2D classes, EM density colored by local resolution, and angular distribution coverage.
Top panel: Micrographs were divided into four batches and classified using 3D classification in RELION with a 60 Å low-pass filtered map of Pol III as a reference. The best class of each batch was retained, batches were combined and refined. Focused classification using a mask on Maf1 yielded one class with improved Maf1 occupancy. Lastly, focused classification using a mask on the stalk-heterotrimer-clamp module separated an open clamp state with poorly resolved Maf1 density from a closed-clamp state with clear separation of the Maf1 β-strands and side-chain density. Bottom panel: FSC curves showing the correlation between independent half maps (left) and the correlation between the sharpened experimental map and a simulated model map (right).
Shown are sequences of loop 1, loop 2 and W319 from distantly related species. Functionally important sites based on mutant studies are boxed in red.
Extended Data Fig. 4 Comparison of the Pol III conformation in apo-Pol III, Maf1-Pol III and the Pol III-PIC.
Maf1-Pol III adopts a similar conformation as the Pol III-PIC (note the distance between the lobe (blue) and clamp head (red)).
Top panel: Structures shown in ribbon representation with α-helices colored in red and β-sheets colored in yellow. Bottom panel: surface representation colored by electrostatic potential from negative (red) to positive (blue) potential.
The phospho-regulatory region of Maf1 is accessible in the Maf1-Pol III structure, whereas the acidic tail emerges in the direction of the DNA binding cleft. The downstream DNA (PDB 6F40) has been superimposed with the Maf1-Pol III structure to help visualization. Maf1 is colored from N-terminus (blue) to C-terminus (red). Putative paths of the disordered regions are indicated, with the internal, phospho-regulated region located away from the Pol III interface, whereas the acidic C-terminal tail projects towards the DNA binding cleft, where it might help to repel nucleic acids.
a, Amino acid sequences of wild-type ScMaf1 and the various ScMaf1 C-terminal mutants are shown from the end of conserved domain C22 through the acidic terminal region (where present). Amino acid sequences from residue 2 through 326 are represented by //. ScMaf1ΔCt terminates at residue 346 while ScMaf1ΔCtSpMAF1 and ScMaf1ΔCtHsMAF1 proteins contain the terminal amino acids from S. pombe (35 residues, colored in blue) and human (45 residues, colored in pink) MAF1 proteins appended to ScMaf1ΔCt. b, The respiratory defect of the maf1Δ::natMX vector only strain, poor growth on glycerol, is rescued by wild-type ScMaf1 and the three ScMaf1 C-terminal variants. Ten-fold serial dilutions of maf1Δ:: natMX strains containing pRS314 vector, pRS314ScMaf1 or pRS314ScMaf1 C-terminal variants grown at 30 and 37 °C on media with glucose (left panels) and glycerol (right panels) as the carbon source. c, Northern analysis of Pol III transcription and repression shows that the C-terminus of ScMaf1 is not required for the Maf1 repression function in yeast. maf1Δ:: natMX strains containing pRS314ScMaf1 or pRS314ScMaf1 C-terminal variants (ScMaf1ΔCt, ScMaf1ΔCtSpMAF1 and ScMaf1ΔCtHsMAF) were treated with rapamycin or drug vehicle for 1 h. The relative level of Pol III transcription is reported by the amount of pre-tRNALeu transcript normalized to U3 snRNA, expressed relative to the untreated wild-type strain and indicated below each lane.
About this article
Cite this article
Vorländer, M.K., Baudin, F., Moir, R.D. et al. Structural basis for RNA polymerase III transcription repression by Maf1. Nat Struct Mol Biol 27, 229–232 (2020). https://doi.org/10.1038/s41594-020-0383-y