Plasmid Replication Control by Antisense RNAs

Abstract

Plasmids are selfish genetic elements that normally constitute a burden for the bacterial host cell. This burden is expected to favor plasmid loss. Therefore, plasmids have evolved mechanisms to control their replication and ensure their stable maintenance. Replication control can be either mediated by iterons or by antisense RNAs. Antisense RNAs work through a negative control circuit. They are constitutively synthesized and metabolically unstable. They act both as a measuring device and a regulator, and regulation occurs by inhibition. Increased plasmid copy numbers lead to increasing antisense-RNA concentrations, which, in turn, result in the inhibition of a function essential for replication. On the other hand, decreased plasmid copy numbers entail decreasing concentrations of the inhibiting antisense RNA, thereby increasing the replication frequency. Inhibition is achieved by a variety of mechanisms, which are discussed in detail. The most trivial case is the inhibition of translation of an essential replication initiator protein (Rep) by blockage of the rep-ribosome binding site. Alternatively, ribosome binding to a leader peptide mRNA whose translation is required for efficient Rep translation can be prevented by antisense-RNA binding. In 2004, translational attenuation was discovered. Antisense-RNA-mediated transcriptional attenuation is another mechanism that has, so far, only been detected in plasmids of Gram-positive bacteria. ColE1, a plasmid that does not need a plasmid-encoded replication initiator protein, uses the inhibition of primer formation. In other cases, antisense RNAs inhibit the formation of an activator pseudoknot that is required for efficient Rep translation.

Genetic control of ColE1 plasmid stability that is independent of plasmid copy number regulation

“…Processing of this R-loop RNA by RNaseH generates a free 3’–OH end that is extended by DNA polymerase I (Pol I), initiating leading-strand synthesis (Itoh and Tomizawa, 1980). Replication initiation is negatively regulated by an antisense RNA mechanism that maintains copy number constant for a given physiological condition (Brantl, 2014; Camps, 2010). The 108 bp antisense RNA (RNAI) is transcribed from a strong promoter located downstream and in the opposite orientation from the promoter for RNAII.…”

“…Plasmid copy number is largely regulated at the level of replication initiation through an antisense RNA mechanism, which maintains copy number at a constant level for a given physiological condition (reviewed in (Brantl, 2014; Camps, 2010; Polisky, 1988)) (Fig. 1).…”

Interplay of regulatory RNAs and mobile genetic elements in enteric pathogens

“…Due to space limitations, we restrict our review to sRNAs for which target genes have been experimentally identified. For the role of regulatory RNA in plasmid maintenance, as well as the recently discovered CRISPR/Cas and BREX systems, we refer to several excellent articles (Brantl, ; Goldfarb et al ., ; Marraffini, ).…”

Plasmid Rolling-Circle Replication

“…This dual mechanism was first discovered in pMV158 and seems to be widespread among plasmids from Gram-positive bacteria, including the RCR plasmids of the pMV158 family (99,102). In pMV158, the transcriptional repressor CopG binds to its own promoter and represses the transcription of the copG-repB operon.…”