A functional hybrid ribosome binding site in tryptophan operon messenger RNA of Escherichia coli

We present evidence that repair of DNA damage induced by decay of incorporated ¹²⁵I after replication of the labeled duplex of Escherichia coli requires the recA⁺ gene function. Furthermore, only about half of the cells survive after label segregation even when that repair function is present. Our results support the possibility that repair of ¹²⁵I decay-induced lesions is asymmetric, being limited to damage initiated in only one of the two strands of the DNA duplex.     

Different half-lives of messenger RNA corresponding to different segments of the tryptophan operon of Escherichia coli

In genetically derepressed strains (trpR⁻) of Escherichia coli which are growing exponentially, messenger RNA regions corresponding to different segments of the trp operon are labeled with different kinetics, suggesting that operator-proximal and distal regions of trp-mRNA have different half-lives. This conclusion was confirmed by direct measurement of trp-mRNA decay; the half-lives for different mRNA regions at 30 °C were found to be 60 seconds for trpE-mRNA, 75 seconds for trpDC-mRNA, and 95 to 115 seconds for trpBA-mRNA. Deletions of genetic segments within the operator-proximal region of the operon reduce the half-life of trp BA-mRNA. Large deletions which place the BA region near the operator reduce the half-life of trpBA-mRNA to values similar to that of trpE-mRNA in the parental strain. Therefore location in the message rather than primary structure appears to determine the half-life of each mRNA region. Several of the internal deletions have a polar effect on the synthesis of the trpB and trpA polypeptides. However, the reduction in trpBA-mRNA half-life does not appear to be due to polarity because trpBA-mRNA half-life is reduced to the same value in three deletion mutants in which there is a sevenfold difference in polarity. These results are compatible with a model of trp-mRNA degradation in which the initial degradative event occurs near the 5′ end of the mRNA molecule and is followed by over-all degradation in the 3′ direction, with random or non-random delays causing an increase in half-life of about 10% per 1000 nucleotides mRNA. Our findings are not compatible with a model of normal degradation in which the entire mRNA molecule is the target for the initial degradative event.     

Nucleotide sequences from messenger RNA transcribed from the operator-proximal portion of the tryptophan operon of Escherichia coli

³²P-labeled messenger RNA transcribed in vivo from the operator-proximal portion of the tryptophan operon of Escherichia coli was purified by DNA/RNA hybridization. The mRNA preparations obtained were subjected to polyaerylaamide gel electrophoresis, and a number of discrete labeled bands were detected. Characterization of the labeled bands and of purified, unbanded mRNA preparations, by partial sequence analysis of the oligonucleotides obtained following T₁ and pancreatic RNase digestion, revealed that the bands represented discrete segments of the trp mRNA molecule. This observation suggests that endonucleolytic cleavage occurs in vivo at specific sites in the mRNA molecule.     

Decay of messenger ribonucleic acid from the lactose operon of Escherichia coli as a function of growth temperature

The inactivation rates of the first, β-galactosidase, and last, transacetylase, messages of the lactose operon of Escherichia coli were measured at different growth temperatures. The inactivation rate of each message appears to increase exponentially with temperature. The rate constant for this increase is almost twice as high for transacetylase message as it is for β-galactosidase message. The inactivation rate is more a direct function of growth temperature than of growth rate. At 15 °C transacetylase message is inactivated about 2.5 times more slowly than is β-galactosidase message. This difference is not paralleled by a different rate of chemical loss of the β-galactosidase message compared to the distal lac mRNA; all parts of the molecule appear to be lost at the same rate. This same pattern is observed in decay of the total mRNA; loss of capacity to direct peptide synthesis (functional inactivation) occurs at variable rates whereas loss of mRNA mass (chemical degradation) seems to occur at a uniform rate.We conclude that each message has a unique target for inactivation with a specifie temperature coefficient of sensitivity, and the inactivation of a message need not be associated with chemical destruction of the molecule.     

Letter: Decay of messenger RNA from the tryptophan operon of Escherichia coli as a function of growth temperature

The temperature coefficients for functional decay and chemical loss of the proximal and distal parts of the polycistronic messenger RNA of the trp operon are similar in the range of growth temperatures between 10 ° and 37 °C.