Pseudoknot-dependent translational coupling in repBA genes of the IncB plasmid pMU720 involves reinitiation.
Replication of the IncB miniplasmid pMU720 requires synthesis of the replication initiator protein, RepA, whose translation is coupled to that of a leader peptide, RepB. The unusual feature of this system is that translational coupling in repBA has to be activated by the formation of a pseudoknot immediately upstream of the repA Shine-Dalgarno sequence. A small antisense RNA, RNAI, controls replication of pMU720 by interacting with repBA mRNA to inhibit expression of repA both directly, by preventing formation of the pseudoknot, and indirectly, by inhibiting translation of repB. The mechanism of translational coupling in repBA was investigated using the specialized ribosome system, which directs a subpopulation of ribosomes that carry an altered anti-Shine-Dalgarno sequence to translate mRNA molecules whose Shine-Dalgarno sequences have been altered to be complementary to the mutant anti-Shine-Dalgarno sequence. Our data indicate that translation of repA involves reinitiation by the ribosome that has terminated translation of repB. The role of the pseudoknot in this process and its effect on the control of copy number in pMU720 are discussed.
Praszkier
J
J
Department of Microbiology and Immunology, The University of Melbourne, Victoria 3010, Australia. judy@ariel.its.unimelb.edu.au
Pittard
A J
AJ
D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D020029:Base Pairing; D001483:Base Sequence; D018387:Codon, Initiator; D018388:Codon, Terminator; D004265:DNA Helicases; D004261:DNA Replication; D004268:DNA-Binding Proteins; D004926:Escherichia coli; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids / Q000235:genetics*; D014176:Protein Biosynthesis; D011506:Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D012329:RNA, Bacterial / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012270:Ribosomes; D015534:Trans-Activators
D001426:Bacterial Proteins; D018387:Codon, Initiator; D018388:Codon, Terminator; D004268:DNA-Binding Proteins; D011506:Proteins; D012329:RNA, Bacterial; D012333:RNA, Messenger; C048817:RepB protein, Bacteria; D015534:Trans-Activators; C045264:replication initiator protein; D004265:DNA Helicases