title: Mechanisms of backtrack recovery by RNA polymerases I and II.
abstract: During DNA transcription, RNA polymerases often adopt inactive backtracked states. Recovery from backtracks can occur by 1D diffusion or cleavage of backtracked RNA, but how polymerases make this choice is unknown. Here, we use single-molecule optical tweezers experiments and stochastic theory to show that the choice of a backtrack recovery mechanism is determined by a kinetic competition between 1D diffusion and RNA cleavage. Notably, RNA polymerase I (Pol I) and Pol II recover from shallow backtracks by 1D diffusion, use RNA cleavage to recover from intermediary depths, and are unable to recover from extensive backtracks. Furthermore, Pol I and Pol II use distinct mechanisms to avoid nonrecoverable backtracking. Pol I is protected by its subunit A12.2, which decreases the rate of 1D diffusion and enables transcript cleavage up to 20 nt. In contrast, Pol II is fully protected through association with the cleavage stimulatory factor TFIIS, which enables rapid recovery from any depth by RNA cleavage. Taken together, we identify distinct backtrack recovery strategies of Pol I and Pol II, shedding light on the evolution of cellular functions of these key enzymes.
authors.affiliation: Biotechnology Center, Technical University Dresden, 01307 Dresden, Germany; Max Planck Institute of Molecular Cell Biology and Genetics, 01307 Dresden, Germany;
authors.forename: Ana
authors.initials: A
authors.lastname: Lisica
authors.affiliation: Department of Molecular Biology, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany;
authors.forename: Christoph
authors.initials: C
authors.lastname: Engel
authors.affiliation: Biotechnology Center, Technical University Dresden, 01307 Dresden, Germany; Max Planck Institute of Molecular Cell Biology and Genetics, 01307 Dresden, Germany;
authors.forename: Marcus
authors.initials: M
authors.lastname: Jahnel
authors.affiliation: Max Planck Institute for the Physics of Complex Systems, 01187 Dresden, Germany; Center for Advancing Electronics Dresden (cfaed), 01062 Dresden, Germany; Grupo Interdisciplinar de Sistemas Complejos (GISC), 28040 Madrid, Spain;
authors.forename: Édgar
authors.initials: É
authors.lastname: Roldán
authors.affiliation: Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, MO 63110.
authors.forename: Eric A
authors.initials: EA
authors.lastname: Galburt
authors.affiliation: Department of Molecular Biology, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany;
authors.forename: Patrick
authors.initials: P
authors.lastname: Cramer
authors.affiliation: Biotechnology Center, Technical University Dresden, 01307 Dresden, Germany; Max Planck Institute of Molecular Cell Biology and Genetics, 01307 Dresden, Germany; Max Planck Institute for the Physics of Complex Systems, 01187 Dresden, Germany; stephan.grill@biotec.tu-dresden.de.
authors.forename: Stephan W
authors.initials: SW
authors.lastname: Grill
mesh_terms: D004058:Diffusion; D008956:Models, Chemical; D009038:Motion; D052898:Optical Tweezers; D011485:Protein Binding; D021122:Protein Subunits; D012318:RNA Polymerase I / Q000737:chemistry / Q000378:metabolism*; D012319:RNA Polymerase II / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012331:RNA, Fungal / Q000096:biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*; D017384:Sequence Deletion; D013269:Stochastic Processes; D013995:Time; D061805:Transcription Elongation, Genetic / Q000502:physiology*; D035602:Transcriptional Elongation Factors / Q000737:chemistry / Q000378:metabolism
chemical_list: D021122:Protein Subunits; D012331:RNA, Fungal; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins; D035602:Transcriptional Elongation Factors; C018648:transcription factor S-II; D012319:RNA Polymerase II; D012318:RNA Polymerase I
keywords: Pol I; Pol II; backtracking; optical tweezers; transcription