Decay of individual Escherichia coli trp messenger RNA molecules is sequentially ordered

Abstract

Two new features of trp mRNA decay were observed by following the metabolism of chains specifically labeled in their 3′-terminal A-gene sequences. Further formation of RNA was shut down with rifampicin and tryptophan, and the decay and size distribution of chains containing labeled A segments were assessed. The 3′-terminal A gene sequences decayed only after a lag of several minutes. During the lag, intermediates in decay could be extracted from cells. They sedimented in sucrose gradients at positions expected for trp operon mRNA lacking successive gene equivalents from the 5′-end. These results eliminate models for mRNA decay in which molecules begin to decay in bulk from their 3′-ends. Quantitative analyses also eliminate a model in which each gene sequence is at jeopardy from the time of its synthesis. Instead the results are consistent with decay proceeding through a mRNA from a random start at or near the 5′-end, with hold-up points at each inter-cistronic border. When distal portions of long species of total Escherichia coli mRNA other than trp were labeled, there was again a lag of several minutes before the labeled sequences broke down to acid-soluble fragments. During the lag, the labeled segments could again be extracted in progressively shorter chains. In principle, similar analyses can determine the mode of decay of other polycistronic mRNA species.