Global colocalization of the histone-like nucleoid-structuring protein (H-NS) with Rho-dependent terminators and genetic interactions between hns and rho suggest that H-NS aids Rho in suppression of antisense transcription. In contrast, a widely studied nusA deletion proposed to compromise Rho-dependent termination had no effect on antisense or sense Rho-dependent terminators in vivo. NusG is required for efficient termination at minority subsets (∼20%) of both antisense and sense Rho-dependent terminators with lower C/G ratio sequences.
Rho binds C-rich unstructured nascent RNA (high C/G ratio) prior to its ATP-dependent. We report that a major function of the Escherichia coli termination factor Rho and its cofactor, NusG, is suppression of ubiquitous antisense transcription genome-wide. Read moreĭespite the prevalence of antisense transcripts in bacterial transcriptomes, little is known about how their synthesis is controlled.
This strategy should have broad utility for illuminating the role of other important protein complexes. Our findings demonstrate that RNAP chemical-genetic interactions provide a general platform for interrogating structure-function relationships in vivo and for identifying physiological trade-offs of mutations, including those relevant for disease and biotechnology. We discover phenotypes for non-essential insertions, show that clustering mutant phenotypes increases their predictive power for drawing functional inferences, and demonstrate that some RNA polymerase mutants both decrease average cell length and prevent killing by cell-wall targeting antibiotics. Here, we conduct a chemical-genetic analysis of a library of RNAP mutants. Numerous studies have interrogated RNAP mechanism, and RNAP mutations drive Escherichia coli adaptation to many health- and industry-relevant environments, yet a paucity of systematic analyses hampers our understanding of the fitness trade-offs from. The multi-subunit bacterial RNA polymerase (RNAP) and its associated regulators carry out transcription and integrate myriad regulatory signals. We discover phenotypes for non-essential insertions, show that clustering mutant phenotypes increases their predictive power for drawing functional inferences, and illuminate a connection between transcription and cell division. Numerous studies have interrogated the inner workings of RNAP, and mutations in genes encoding RNAP drive adaptation of Escherichia coli to many health- and industry-relevant environments, yet a paucity of systematic analyses has hampered our understanding of the fitness benefits and trade-offs from altering RNAP function.
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