THE MECHANISM OF TRANSLATIONAL COUPLING IN ESCHERICHIA-COLI - HIGHER-ORDER STRUCTURE IN THE ATPHA MESSENGER-RNA ACTS AS A CONFORMATIONAL SWITCH REGULATING THE ACCESS OF DE-NOVO INITIATING RIBOSOMES

Bacterial genes are commonly transcribed to form polycistronic mRNAs b earing reading frames whose respective translational efficiencies are not independently determined. As in many bacterial operons, expression of the atp genes of Escherichia coli is strongly influenced by transl ational coupling. The gene pair atpHA is tightly coupled, whereby atpA is translated at least three times more efficiently than atpH. Howeve r, there is no fixed stoichiometry of coupling: mutations in atpH lead to increases in the translation ratio (atpA/ atpH) of up to approxima tely 40-fold. We have demonstrated that secondary structure sequesteri ng the atpA translational initiation region (TIR) is important to the coupling mechanism in that it inhibits de novo translational initiatio n at the atpA start codon. Genetic and structural analyses indicate th at this inhibitory structure can be induced to refold into a less inhi bitory con formation either by introducing two single-base substitutio ns or as a result of ribosomes translating atpH. We propose a model in which the secondary structure of the atpA TIR acts analogously to a ' 'gating device'' in that it restricts de novo ribosomal initiation unt il it is ''switched'' into a more open conformation. This contrasts wi th the function of a stem-loop structure located immediately downstrea m of atpA and upstream of the Shine-Dalgarno region of atpG, which was found to inhibit translation, but not to mediate tight coupling. Resu lts obtained using the ''specialized'' ribosome system of Hui and de B oer ((1987) Proc. Natl. Acad. Sci. U. S. A. 84, 4762-4766) indicate th at primarily ribosomes reinitiating after termination on atpH are resp onsible for inducing refolding of the atpA TIR. The principle of alter native mRNA conformations with different functional properties embodie d in the model presented here can only be fulfilled by certain types o f structure. It is likely to operate in several steps of prokaryotic g ene expression, underlying a range of regulatory events including tran scriptional attenuation and translational activation.