6S RNA is a bacterial transcriptional regulator,which accumulates during 
stationary phase and inhibits transcription from many promoters due to stable 
association with σ 70 -containing RNA polymerase. This inhibitory RNA polymerase 
∼ 6S RNA complex dissociates during nutritional upshift, when cells undergo 
outgrowth from stationary phase, releasing active RNA polymerase ready for 
transcription. The release reaction depends on a characteristic property of 6S 
RNAs, namely to act as template for the de novo synthesis of small RNAs, termed 
pRNAs.Here, we used limited hydrolysis with structure-specific RNases and 
in-line probing of isolated 6S RNA and 6SRNA ∼ pRNA complexes to investigate the 
molecular details leading to the release reaction. Our results indicate that 
pRNA transcription induces the refolding of the 6S RNA secondary structure by 
disrupting part of the closing stem(conserved sequence regions CRI and CRIV) and 
formation of a new hairpin (conserved sequence regions CRIII and CRIV). 
Comparison of the dimethylsulfate modification pattern of 6S RNA in living cells 
at stationary growth and during outgrowth confirmed the conformational change 
observed in vitro. Based on our results, a model describing the individual steps 
of the release reaction is presented.