PMID:20354151
| Citation |
Shephard, L, Dobson, N and Unrau, PJ (2010) Binding and release of the 6S transcriptional control RNA. RNA 16:885-92 |
|---|---|
| Abstract |
6S RNA is an important noncoding RNA that regulates eubacterial transcription. In Escherichia coli this RNA binds to the sigma(70) RNA polymerase holoenzyme and is released by the synthesis of a short product RNA. In order to determine how binding and release are controlled by the 6S RNA sequence, we used in vitro selection to screen a high diversity library containing approximately 4 x 10(12) sequences for functional 6S RNA variants. Residues critical for binding were found to be located in a "-35" region upstream of the 6S RNA transcription bubble mimic structure. Mutating these phylogenetically conserved residues invariably led to decreases in binding and removing them abolished binding, implicating these nucleotides in a biologically important interaction with the Esigma(70) complex. Interestingly, mutation of phylogenetically conserved "-10" residues that were also upstream of the site of pRNA synthesis was found to influence 6S RNA release rates in addition to modulating -35 binding. These results indicate how 6S RNA -35 binding to sigma(70) RNA polymerase holoenzyme can regulate expression from "strong" and "weak" -35 DNA promoters and suggest that 6S RNA release rates have been fine tuned over evolutionary time so as to correctly regulate cellular levels of transcription. |
| Links |
PubMed PMC2856883 Online version:10.1261/rna.2036210 |
| Keywords |
Base Sequence; Binding Sites/genetics; Consensus Sequence; Conserved Sequence; DNA-Directed RNA Polymerases/metabolism; Escherichia coli K12/genetics; Escherichia coli K12/metabolism; Evolution, Molecular; Gene Library; Kinetics; Models, Biological; Models, Molecular; Molecular Sequence Data; Mutagenesis, Site-Directed; Nucleic Acid Conformation; Phylogeny; Promoter Regions, Genetic; RNA, Bacterial/chemistry; RNA, Bacterial/genetics; RNA, Bacterial/metabolism; Sequence Deletion; Sigma Factor/metabolism |
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