Expression of the replication protein Arp of phasyl shows dual regulation by an antisense promoter.

Phasyl is the smallest naturally occurring replicon found so far in Escherichia coli. It encodes a protein which is essential for autonomous replication (Arp). The transcriptional start of the arp gene was mapped. A strong antisense promoter was found in close proximity to the arp promoter. The inactivation of this promoter led in cis to a strong increase of the transcription of the arp gene and to the inactivation of autonomous replication of phasyl. The product of the antisense promoter is an 83 nt RNA molecule, which is not translated. The antisense RNA led in trans to the inhibition of the translation of the arp mRNA, presumably mediated by the formation of an RNA-RNA hybrid in which the Shine-Dalgarno sequence of the arp transcript is sequestered. The expression of the arp gene is thus controlled by two negatively acting mechanisms: it is subject to a transcriptional control in cis exerted by the antisense promoter and to a translational control in trans mediated by the antisense RNA. Inactivation of one mechanism of control cannot be compensated by the remaining one.