Antisense RNA and DNA in Escherichia coli

Abstract

Antisense RNA (asRNA) has been shown to be utilized effectively in Escherichia coli as a natural regulatory factor to control a variety of important biological processes. Among them are: plasmid replication, IS10 transposition, osmoregulation of ompF gene expression, autoregulation of cAMP-receptor protein synthesis, cell-killing function through activity of hok homologs, cell division, and P22 and λ phage development. In most cases, asRNAs are small untranslatable transcripts that pair with a target RNA sequence and exert a negative control by not allowing the normal interaction of target RNA with other nucleic acids or protein factors, or lead to the increased rate of degradation by RNase specific for double-stranded RNA. Some of the asRNAs are synthesized in significant quantities and have a high turnover rate, whereas others, which mostly regulate transposon and phage actions, are expressed at low levels, but are significantly stable. Members of the asRNA family may be divided into two major groups: asRNAs that are transcribed from the same loci as the target genes and asRNAs whose genes locate at genetic loci different from the target genes acting as transcoded regulators. AsRNAs belonging to the first group are completely complementary to their target RNA, whereas asRNAs from the second group usually form only partially complementary hybrids with their target sequences. Retroelements found in gram-negative bacteria allow a novel method of in vivo synthesis of specific oligonucleotides, providing an efficient system for the regulation of gene expression in E. coli. It is based on the complementarity of single-stranded DNA (asDNA) possessing antisense sequences to specific genes. The principle of the method is to express asDNA as a part of msDNA, which are produced in E. coli by bacterial reverse transcriptase. This chapter describes approaches using both antisense RNA and antisense DNA for the regulation of gene expression in E. coli.