Characterizing a new role of TRAP in the transcription attenuation mechanism of the Bacillus subtilis Trp operon
Potter, Kristine Dara
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In Bacillus subtilis, the trpEDCFBA operon is regulated by a transcription attenuation mechanism controlled by the trp RNA-binding attenuation protein (TRAP). When TRAP is activated by tryptophan, it binds to 11 (G/U)AG repeats in the trp leader transcript and prevents formation of an antiterminator, which allows formation of an intrinsic terminator (attenuator). Previously, formation of the attenuator RNA structure was believed to be solely responsible for signaling RNA polymerase (RNAP) to halt transcription. However, this study shows that substitutions that prevent formation of the antiterminator do not result in efficient transcription termination. The attenuator requires the presence of TRAP bound to the nascent RNA to cause efficient transcription termination, suggesting TRAP has an additional more active role in causing termination at the attenuator. Two characteristic features of intrinsic terminators, a GC-rich stem and uninterrupted U-stretch, are instead an AU-rich hairpin and interrupted U-stretch in the trp attenuator. These deviations from consensus result in the requirement of TRAP both in vitro and in vivo for efficient termination. In one model for intrinsic termination, hairpin formation in the nascent RNA causes RNAP to translocate downstream on the DNA template in the absence of RNA synthesis, leading to transcript release. This model is supported by studies of the energetics of transcription termination, which indicate that forward movement of RNAP is required when the U-tract after the hairpin is interrupted, as occurs with the trp attenuator. This study also shows that transcript release at the trp attenuator requires forward movement of RNAP and that the additional role of TRAP is to assist this movement to cause efficient transcription termination.