Messenger RNA secondary structure and translational coupling in the Escherichia coli operon encoding translation initiation factor IF3 and the ribosomal proteins, L35 and L20.
The Escherichia coli infC-rpmI-rplT operon encodes translation initiation factor IF3 and the ribosomal proteins, L35 and L20, respectively. The expression of the last cistron (rplT) has been shown to be negatively regulated at a post-transcriptional level by its own product, L20, which acts at an internal operator located within infC. The present work shows that L20 directly represses the expression of rpmI, and indirectly that of rplT, via translational coupling with rpmI. Deletions and an inversion of the coding region of rpmI, suggest an mRNA secondary structure forming between sequences within rpmI and the translation initiation site of rplT. To verify the existence of this structure, detailed analyses were performed using chemical and enzymatic probes. Also, mutants that uncoupled rplT expression from that of rpmI, were isolated. The mutations fall at positions that would base-pair in the secondary structure. Our model is that L20 binds to its operator within infC and represses the translation of rpmI. When the rpmI mRNA is not translated, it can base-pair with the ribosomal binding site of rplT, sequestering it, and abolishing rplT expression. If the rpmI mRNA is translated, i.e. covered by ribosomes, the inhibitory structure cannot form leaving the translation initiation site of rplT free for ribosomal binding and for full expression. Although translational coupling in ribosomal protein operons has been suspected to be due to the formation of secondary structures that sequester internal ribosomal binding sites, this is the first time that such a structure has been shown to exist.
Lesage
P
P
Institut de Biologie Physico-Chimique, Paris, France.
Chiaruttini
C
C
Graffe
M
M
Dondon
J
J
Milet
M
M
Springer
M
M
D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009876:Operon; D010448:Peptide Initiation Factors / Q000235:genetics*; D039665:Prokaryotic Initiation Factor-3; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics*; D015183:Restriction Mapping; D012269:Ribosomal Proteins / Q000235:genetics*; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; C040045:L20 ribosomal protein, bacteria; D010448:Peptide Initiation Factors; D039665:Prokaryotic Initiation Factor-3; D012333:RNA, Messenger; D012269:Ribosomal Proteins; C489279:ribosomal protein L35, E coli; C488992:rplT protein, E coli