title: Occlusion of the Ribosome Binding Site Connects the Translational Initiation Frequency, mRNA Stability and Premature Transcription Termination.
abstract: Protein production is controlled by ribosome binding to the messenger RNA (mRNA), quantified in part by the binding affinity between the ribosome and the ribosome binding sequence on the mRNA. Using theE. coli lacoperon as model, Ringquist et al. (1992) found a more than 1,000-fold difference in protein yield when varying the Shine-Dalgarno sequence and its distance to the translation start site. Their proposed model accounted for this large variation by only a variation in the binding affinity and the subsequent initiation rate. Here we demonstrate that the decrease in protein yield with weaker ribosome binding sites in addition is caused by a decreased mRNA stability, and by an increased rate of premature transcription termination. Using different ribosome binding site sequences of theE. coli lacZgene, we found that an approximately 100-fold span in protein expression could be subdivided into three mechanisms that each affected expression 3- to 6-fold. Our experiments is consistent with a two-step ribosome initiation model, in which occlusion of the initial part of the mRNA by a ribosome simultaneously protects the mRNA from both premature transcription termination and degradation: The premature termination we suggest is coupled to the absence of occlusion that allows binding of transcription termination factor, possibly Rho. The mRNA stability is explained by occlusion that prevents binding of the degrading enzymes. In our proposed scenario, a mRNA with lower translation initiation rate would at the same time be "hit" by an increased premature termination and a shorter life-time. Our model further suggests that the transcription from most if not all natural promoters is substantially influenced by premature termination.
authors.affiliation: Department of Biology, University of Copenhagen Copenhagen, Denmark.
authors.forename: Mette
authors.initials: M
authors.lastname: Eriksen
authors.affiliation: Center for Models of Life, Niels Bohr Institute, University of Copenhagen Copenhagen, Denmark.
authors.forename: Kim
authors.initials: K
authors.lastname: Sneppen
authors.affiliation: Department of Biology, University of CopenhagenCopenhagen, Denmark; Center for Models of Life, Niels Bohr Institute, University of CopenhagenCopenhagen, Denmark.
authors.forename: Steen
authors.initials: S
authors.lastname: Pedersen
authors.affiliation: Center for Models of Life, Niels Bohr Institute, University of Copenhagen Copenhagen, Denmark.
authors.forename: Namiko
authors.initials: N
authors.lastname: Mitarai
keywords: gene expression; mRNA stability; modeling; occlusion time; transcription termination; translation initiation frequency