The long-term goal of this research project is to understand the basic principles that govern mRNA degradation, a process that has long been known to play a key role in controlling gene expression in all organisms. In Escherichia coli and related bacterial species, the ribonuclease with the greatest influence on mRNA lifetimes is RNase E, an endonuclease with a strong preference for decay intermediates that bear a single phosphate at the 5? terminus. Recent evidence indicates that RNase E locates its cleavage sites in monophosphorylated RNA by scanning linearly downstream from the 5? end along RNA segments that are single-stranded. The immediate objectives of this research project are to elucidate the mechanism of RNase E scanning, the enzyme characteristics that make it possible, the features of RNA that facilitate or impede it, and the impact of this process on gene expression. Achieving these objectives will require the use of standard molecular biological, biochemical, and genetic methods as well as single-molecule FRET. The knowledge gained from these studies will provide fundamental insights into a novel aspect of gene regulation that is likely to be important for bacterial pathogenesis.
The proposed research will address a key aspect of the mechanism by which messenger RNA is degraded in bacterial cells. The knowledge thereby acquired is expected to be of value in understanding the regulatory processes that govern bacterial pathogenesis and in maximizing bacterial production of medically useful proteins. In addition, the methods and concepts developed in the course of these studies are likely to be useful for elucidating how messenger RNA degradation helps to ensure proper levels of gene expression in healthy human cells.
| Luciano, Daniel J; Belasco, Joel G (2018) Analysis of RNA 5' ends: Phosphate enumeration and cap characterization. Methods : |
