The Optimal Eukaryotic Signal for Translation Initiation From Non-AUG Codons, Present Upstream of Bacteriophage Lambda P Cistron, is Inactive In Escherichia … (original) (raw)
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Cellular & molecular biology letters, 2003
Expression of the replication genes of bacteriophage lambda, O and P, is believed to be translationally coupled. However, it was previously noted that, under conditions of amino acid starvation, when O is not synthesized, P continues to be expressed at a relatively high level. The results presented in this report, contrary to the previously presented hypothesis, suggest that an AGACUGGAU sequence (an optimal context for translation initiation from non-AUG codons in eukaryotes, and present upstream the P cistron) is inactive in Escherichia coli. Comparative sequence analysis confirms that such a signal is unlikely to be important for P synthesis. Instead, a weak Shine-Dalgarno sequence may be present upstream the P cistron, and be active in the absence of O gene expression.
Cellular & Molecular Biology Letters
Expression of the replication genes of bacteriophage λ, O and P, is believed to be translationally coupled. However, it was previously noted that, under conditions of amino acid starvation, when O is not synthesized, P continues to be expressed at a relatively high level. The results presented in this report, contrary to the previously presented hypothesis, suggest that an AGACUGGAU sequence (an optimal context for translation initiation from non-AUG codons in eukaryotes, and present upstream the P cistron) is inactive in Escherichia coli. Comparative sequence analysis confirms that such a signal is unlikely to be important for P synthesis. Instead, a weak Shine-Dalgarno sequence may be present upstream the P cistron, and be active in the absence of O gene expression.
Proceedings of the National Academy of Sciences, 1988
The late operon of bacteriophage A contains the genes encoding the morphogenetic proteins of the phage. These genes are transcribed equally from the single late promoter. Although the functional half-lives of the mRNA for the various genes of this operon vary <2-fold, their relative rates of expression have been shown to vary by nearly 1000fold. This variation could result from differing rates of translation initiation, from overlapping upstream translation, or from differential elongation rates due to the presence of codons for which the corresponding tRNAs are rare. To distinguish between these possibilities, we have cloned sequences surrounding the initiator codons of several of these genes and measured their ability to drive synthesis of hybrid A-agalactosidase proteins. The rates of expression of the hybrid genes thus produced correlate very well with the natural rates of expression of the corresponding phage genes, suggesting that the rate of initiation of translation controls the relative expression rates of these genes.
Journal of Bacteriology
The bacteriophage A cIl gene product regulates the lysogenic pathway. The cIl gene is located in the leftward operon, which is transcribed from the PL promoter. We have previously shown (S. Altuvia and A. B. Oppenheim, J. Bacteriol. 167:415-419, 1986) that mutations that show elevated expression lie within the clll coding sequence. We isolated mutants that show decreased CIII activity. AUl the mutations were found to cause
Translation initiation in Escherichia coli: sequences within the ribosome-binding site
Molecular Microbiology, 1992
The translational roles of the Shine-Dalgarno sequence, the initiation codon, the space between them, and the second codon have been studied. The Shine-Dalgarno sequence UAAGGAGG initiated translation roughly four times more efficiently than did the shorter AAGGA sequence. Each Shine-Dalgarno sequence required a minimum distance to the initiation codon in order to drive translation; spacing, however, could be rather long. Initiation at AUG was more efficient than at GUG or UUG at each spacing examined; initiation at GUG was only slightly better than UUG. Translation was also affected by residues 3' to the initiation codon. The second codon can influence the rate of initiation, with the magnitude depending on the initiation codon. The data are consistent with a simple kinetic model in which a variety of rate constants contribute to the process of translation initiation.
Inhibition of Escherichia coli protein synthesis by abortive translation of phage λ minigenes
Journal of Molecular Biology, 1997
Escherichia coli mutants defective in peptidyl-tRNA hydrolase activity are unable to maintain bacteriophage lambda vegetative growth. Phage mutants, named bar, overcome the host limitation to support viral growth. Multicopy expression of lambda wild-type bar regions is deleterious to hydrolase-defective cells because it provokes arrest of protein synthesis. We noticed that the bar regions include minigenes whose transcripts would contain a Shine-Dalgarno-like sequence appropriately spaced for translation from a two codon open reading frame. To investigate the mechanism of bar inhibition, we asked if transcripts of the barI region function as mRNAs in their ribosomal interactions. We found that bar-containing RNA associates with ribosomes, forms ternary initiation complexes, yields a toeprint signal, and can be removed from ribosomes by run-off translation, as authentic mRNA. Since bar-containing RNA has the properties of a messenger, we propose that its translation leads to drop-off and accumulation of peptidyl-tRNA in pth-defective cells. Starvation of the tRNA(s) sequestered in pepidyl-tRNA(s) eventually causes inhibition of protein synthesis.
FEBS Letters, 2000
In Eubacteria, de novo translation of some internal cistrons may be inefficient or impossible unless the 5P P neighboring cistron is also translated (translational coupling). Translation reinitiation is an extreme case of translational coupling in which translation of a message depends entirely on the presence of a nearby terminating ribosome. In this work, the characteristics of mRNA cis-elements inducing the reinitiation process in Escherichia coli have been investigated using a combinatorial approach. A number of novel translational reinitiation sequences (TRSs) were thus identified, which show a wide range of reinitiation activities fully dependent on a translational coupling event and unrelated to the presence/absence of secondary structure or mRNA stability. Moreover, some of the isolated TRSs are similar to intercistronic sequences present in the E. coli genome.
Molecular Microbiology, 1998
We determined the in vivo translational efficiency of 'unleadered' lacZ compared with a conventionally leadered lacZ with and without a Shine-Dalgarno (SD) sequence in Escherichia coli and found that changing the SD sequence of leadered lacZ from the consensus 5Ј-AGGA-3Ј to 5Ј-UUUU-3Ј results in a 15-fold reduction in translational efficiency; however, removing the leader altogether results in only a twofold reduction. An increase in translation coincident with the removal of the leader lacking a SD sequence suggests the existence of stronger or novel translational signals within the coding sequence in the absence of the leader. We examined, therefore, a change in the translational signals provided by altering the AUG initiation codon to other naturally occurring initiation codons (GUG, UUG, CUG) in the presence and absence of a leader and find that mRNAs lacking leader sequences are dependent upon an AUG initiation codon, whereas leadered mRNAs are not. This suggests that mRNAs lacking leader sequences are either more dependent on perfect codon-anticodon complementarity or require an AUG initiation codon in a sequence-specific manner to form productive initiation complexes. A mutant initiator tRNA with compensating anticodon mutations restored expression of leadered, but not unleadered, mRNAs with UAG start codons, indicating that codon-anticodon complementarity was insufficient for the translation of mRNA lacking leader sequences. These data suggest that a cognate AUG initiation codon specifically serves as a stronger and different translational signal in the absence of an untranslated leader.
Translational signals of a major head protein gene of bacteriophage lambda
MGG Molecular & General Genetics, 1988
The D gene of bacteriophage 2 which codes for a major head protein is expressed at a high level during lytic growth. We have constructed a set of D-lacZ gene fusions in order to examine the factors determining the high efficiency of the D translational initiation signals. It was found that an integral sequence, 300 bp long and upstream of the ATG initiation codon, is required for maximal protein synthesis.