Biochimie (1991) 73, 1573-1578 1573 © Socirt6 frangaise de biochimie et biologic molrculaire / Elsevier, Paris Ribosomal RNA and peptidyl-tRNA hydrolase: a peptide chain termination model for iambda bar RNA inhibition EJ Murgola l, G Guameros 2 t Department of Molecular Genetics, The University of Texas MD Anderson Cancer Center, 1515 Holcombe Boulevard, Houston, TX 77030 USA; 2Departmento de Genetica y Biologia Molecular, Centro de lnvestigacion y de Estudios Avanzados del IPN, Apostado Postal 14-740, Mexico DF 07000 (Received l November 1991; accepted 6 November 1991) Summary -- We propose here a model to explain the inhibition of bacteriophage lambda (~.) vegetative growth and the killing of E coli cells defective in peptidyl-tRNA hydrolase (Pth) by ~, bar RNA. The model suggests that bar RNA, which contains a character- istic UGA triplet, base-pairs in an anti-parallel fashion with the t 199-1205 region of E coli 16S rRNA. In doing so, it prevents the required functioning of that region of 16S rRNA in UGA-specific peptide chain termination. Pth is implicated in peptide chain termin- ation because a defect in Pth is required for the achievement of the bar RNA inhibitory effects. We make certain predictions that flow from the model, predictions involving suppression of nonsense mutations, and present preliminary experimental results that demon- strate the fulfillment of those predictions. codon recognition / 16S rRNA / nonsense suppression / bar/16S synergism / antisense RNA target Introduction It was previously observed that, in peptidyl-tRNA hydrolase (Pth)-defective cells, transcription of the bar region of bacteriophage lambda (~.) led to inhi- bition of ~, vegetative growth. Furthermore, when bar + was cloned into a high copy plasmid, the presence of the bar + ~lasmid was lethal for Pth- (but not Pth +) cells. To explain these and related observations, we propose here a model that points to a ribosomal RNA nucleotide sequence ~ a target for anti-parallel base pairing with ~, bar RNA. The model also implicates peptidyl-tRNA hydrolase in the normal peptide chain termination process. We state a few basic predictions of the model and present preliminary evidence for the fulfilment of those predictions. Finally, we discuss some ramifications of the model. The phenomenon of lambda bar inhibition The rap mutant of E coli is unable to support the growth of bacteriophage ~ [1, 2]. Phage mutations that overcome the rap effect have been mapped to different sites in the phage genome, and two of those sites were designated barI and barII [3]. Plasmids containing wild-type ~, bar DNA segments kill rap mutant cells, but plasmids with mutant bar regions do not [4]. The plasmid phenomenon closely parallels the phage behavior (table I)0 but during phage infection, cell lethality is not required for inhibition of ~, growth (D Vazquez and G Guameros, unpublished results). It- _L1- rr~ ..... . r~--¢':~-- ~4~ .... '|,¢,i ..... L--u ]I~.T A is necessaa-y uuul lll~Llli~gu,~lllJtlVO'll lull WIIId--L~/laIK; OUt l.JIN/'~k to prevent growth of ~, and for bar ~ plasmid lethality to E coli [3, 4]. The magnitude of transcription through barI and barlI correlates with the degree of Rap inhibition. After transcription induction, Bar* RNA accumulates much more rapidly than Bar- RNA and exhibits greater stability (3.5 min half-life versus less than 1 min) in Rap- bacteria [6]. Table I. Summary of k Bar effects (adapted from [5]). E coli ~, vegetative Growth of strain growth at 39°C transformed cellsa at 40°C bar + A bar- p bar + p bar- pth + +b + + + pth-C _b + _ + abar+ and bar- regions of k were cloned under the tran- scriptional control of E coli pGal promoter, b+, represents proper formation of plaques: -, absence of plaques. CThe pth- was the thermosensitive mutant pth(ts). This strain grows at 40°C, although poorly.