title: Competitive folding of RNA structures at a termination-antitermination site.
abstract: Antitermination is a regulatory process based on the competitive folding of terminator-antiterminator structures that can form in the leader region of nascent transcripts. In the case of theBacillus subtilis licSgene involved in β-glucosides utilization, the binding of the antitermination protein LicT to a short RNA hairpin (RAT) prevents the formation of an overlapping terminator and thereby allows transcription to proceed. Here, we monitored in vitro the competition between termination and antitermination by combining bulk and single-molecule fluorescence-based assays using labeled RNA oligonucleotide constructs of increasing length that mimic the progressive transcription of the terminator invading the antiterminator hairpin. Although high affinity binding is abolished as soon as the antiterminator basal stem is disrupted by the invading terminator, LicT can still bind and promote closing of the partially unfolded RAT hairpin. However, binding no longer occurs once the antiterminator structure has been disrupted by the full-length terminator. Based on these findings, we propose a kinetic competition model for the sequential events taking place at the termination-antitermination site, where LicT needs to capture its RAT target before completion of the terminator to remain tightly bound during RNAP pausing, before finally dissociating irreversibly from the elongatedlicStranscript.
authors.affiliation: CNRS UMR5048, Centre de Biochimie Structurale, 34090 Montpellier, France.|INSERM U1054, 34090 Montpellier, France.|Université de Montpellier, 34090 Montpellier, France.
authors.forename: Soraya
authors.initials: S
authors.lastname: Ait-Bara
authors.affiliation: CNRS UMR5048, Centre de Biochimie Structurale, 34090 Montpellier, France.|INSERM U1054, 34090 Montpellier, France.|Université de Montpellier, 34090 Montpellier, France.
authors.forename: Caroline
authors.initials: C
authors.lastname: Clerté
authors.affiliation: CNRS UMR5048, Centre de Biochimie Structurale, 34090 Montpellier, France.|INSERM U1054, 34090 Montpellier, France.|Université de Montpellier, 34090 Montpellier, France.|INRA, departement MICA, 78352 Jouy-en-Josas, France.
authors.forename: Nathalie
authors.initials: N
authors.lastname: Declerck
authors.affiliation: CNRS UMR5048, Centre de Biochimie Structurale, 34090 Montpellier, France.|INSERM U1054, 34090 Montpellier, France.|Université de Montpellier, 34090 Montpellier, France.
authors.forename: Emmanuel
authors.initials: E
authors.lastname: Margeat
mesh_terms: D020121:5' Untranslated Regions; D001426:Bacterial Proteins / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D009690:Nucleic Acid Conformation; D059370:RNA Folding; D016601:RNA-Binding Proteins / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D061806:Transcription Termination, Genetic; D014158:Transcription, Genetic
chemical_list: D020121:5' Untranslated Regions; D001426:Bacterial Proteins; C098955:LicT protein, Bacteria; D016601:RNA-Binding Proteins; D014157:Transcription Factors; C064872:antiterminator proteins, Bacteria
keywords: FRET; antitermination; fluorescence; single molecule; transcription