title: High-Resolution Phenotypic Landscape of the RNA Polymerase II Trigger Loop.
abstract: The active sites of multisubunit RNA polymerases have a "trigger loop" (TL) that multitasks in substrate selection, catalysis, and translocation. To dissect the Saccharomyces cerevisiae RNA polymerase II TL at individual-residue resolution, we quantitatively phenotyped nearly all TL single variants en masse. Three mutant classes, revealed by phenotypes linked to transcription defects or various stresses, have distinct distributions among TL residues. We find that mutations disrupting an intra-TL hydrophobic pocket, proposed to provide a mechanism for substrate-triggered TL folding through destabilization of a catalytically inactive TL state, confer phenotypes consistent with pocket disruption and increased catalysis. Furthermore, allele-specific genetic interactions among TL and TL-proximal domain residues support the contribution of the funnel and bridge helices (BH) to TL dynamics. Our structural genetics approach incorporates structural and phenotypic data for high-resolution dissection of transcription mechanisms and their evolution, and is readily applicable to other essential yeast proteins.
authors.affiliation: Department of Biochemistry & Biophysics, Texas A&M University, College Station, Texas.
authors.forename: Chenxi
authors.identifier: http://orcid.org/0000-0002-0314-5307
authors.initials: C
authors.lastname: Qiu
authors.affiliation: Mississippi State University, Starkville, Mississippi.
authors.forename: Olivia C
authors.initials: OC
authors.lastname: Erinne
authors.affiliation: Department of Biochemistry & Biophysics, Texas A&M University, College Station, Texas.|Yale Cardiovascular Research Center, Dept. of Internal Medicine, Yale University School of Medicine, New Haven, Connecticut.
authors.forename: Jui M
authors.initials: JM
authors.lastname: Dave
authors.affiliation: Department of Biochemistry & Biophysics, Texas A&M University, College Station, Texas.|Department of Veterinary Pathobiology, Texas A&M University, College Station, Texas.
authors.forename: Ping
authors.initials: P
authors.lastname: Cui
authors.affiliation: Department of Biochemistry & Biophysics, Texas A&M University, College Station, Texas.|Roche Nimblegen, Madison, Wisconsin.
authors.forename: Huiyan
authors.initials: H
authors.lastname: Jin
authors.affiliation: Department of Biochemistry & Biophysics, Texas A&M University, College Station, Texas.|Shire Pharmaceutical, Boston, Massachusetts.
authors.forename: Nandhini
authors.initials: N
authors.lastname: Muthukrishnan
authors.affiliation: Department of Biochemistry & Biophysics, Texas A&M University, College Station, Texas.|Department of Veterinary Pathobiology, Texas A&M University, College Station, Texas.
authors.forename: Leung K
authors.initials: LK
authors.lastname: Tang
authors.affiliation: Department of Biochemistry & Biophysics, Texas A&M University, College Station, Texas.
authors.forename: Sabareesh Ganesh
authors.identifier: http://orcid.org/0000-0002-5551-6683
authors.initials: SG
authors.lastname: Babu
authors.affiliation: Department of Biochemistry & Biophysics, Texas A&M University, College Station, Texas.|College of Medicine, Texas A&M Health Science Center, College Station, Texas.
authors.forename: Kenny C
authors.initials: KC
authors.lastname: Lam
authors.affiliation: Department of Biochemistry & Biophysics, Texas A&M University, College Station, Texas.|Department of Biochemistry & Molecular Biology, Baylor College of Medicine, Houston, Texas.
authors.forename: Paul J
authors.identifier: http://orcid.org/0000-0002-9795-7647
authors.initials: PJ
authors.lastname: Vandeventer
authors.affiliation: MorphoSys AG, Planegg, Germany.
authors.forename: Ralf
authors.initials: R
authors.lastname: Strohner
authors.affiliation: MorphoSys AG, Planegg, Germany.
authors.forename: Jan
authors.initials: J
authors.lastname: Van den Brulle
authors.affiliation: Department of Biochemistry & Biophysics, Texas A&M University, College Station, Texas.|Department of Computer Science and Engineering, Texas A&M University, College Station, Texas.
authors.forename: Sing-Hoi
authors.initials: SH
authors.lastname: Sze
authors.affiliation: Department of Biochemistry & Biophysics, Texas A&M University, College Station, Texas.
authors.forename: Craig D
authors.initials: CD
authors.lastname: Kaplan
mesh_terms: D000483:Alleles; D002384:Catalysis; D020134:Catalytic Domain / Q000235:genetics; D018360:Crystallography, X-Ray; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics*; D009154:Mutation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D021381:Protein Transport / Q000235:genetics; D012319:RNA Polymerase II / Q000737:chemistry / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D013379:Substrate Specificity; D014158:Transcription, Genetic
chemical_list: D050505:Mutant Proteins; D012319:RNA Polymerase II