SsrA tagging of Escherichia coli SecM at its translation arrest sequence.
SecM is expressed from the secM-secA operon and activates the expression of secA in response to secretion defects. The 3'-end of secM encodes an "arrest sequence," which can interact with the ribosomal exit tunnel, preventing complete secM translation under secretion-defective conditions. In a cis-acting manner, ribosome stalling enhances secA translation. Pro(166) is the last residue incorporated when SecM elongation is arrested. We report that secretion deficiencies lead to SsrA tagging of SecM after Pro(166), Gly(165), and likely Arg(163). Northern blot analysis revealed the presence of a truncated secM transcript, likely issued from a secM-secA cleavage. The level of secM transcripts was decreased either when secM translation was totally prevented or when Pro(166) was mutated. However, the accumulation of a truncated secM transcript required secM translation and was prevented when the SecM arrest sequence was inactivated by a point mutation changing Pro(166) to Ala. We suggest that ribosome pausing at the site encoding the arrest sequence is required for formation of the truncated secM mRNA. SsrA tagging affected neither the presence of the secM mRNA nor secA expression, even under translocation-defective conditions. It is therefore likely that SsrA tagging of SecM occurs only after cleavage of secM-secA mRNA within the secM open reading frame encoding the SecM arrest sequence. Accumulation of transcripts expressing arrested SecM generated growth inhibition that was alleviated by the SsrA tagging system. Therefore, SsrA tagging of SecM would rescue ribosomes to avoid excessive jamming of the translation apparatus on stop-less secM mRNA.
Collier
Justine
J
Laboratoire des Réseaux de Régulations et Biogenèse de l'Enveloppe Bactérienne, Institut de Génétique et Microbiologie, Université Paris-Sud, CNRS, UMR8621, Bātiment 400, F-91405 Orsay Cedex, France.
Bohn
Chantal
C
Bouloc
Philippe
P
D000251:Adenosine Triphosphatases / Q000235:genetics; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics; D015152:Blotting, Northern; D015153:Blotting, Western; D003593:Cytoplasm / Q000737:chemistry; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics* / Q000648:ultrastructure; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D026901:Membrane Transport Proteins / Q000235:genetics; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016366:Open Reading Frames; D010441:Peptide Chain Elongation, Translational; D017354:Point Mutation; D014176:Protein Biosynthesis; D012329:RNA, Bacterial / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D012343:RNA, Transfer / Q000032:analysis / Q000235:genetics; D012270:Ribosomes / Q000378:metabolism; D000069816:SEC Translocation Channels; D000081416:SecA Proteins; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D012329:RNA, Bacterial; D012333:RNA, Messenger; D000069816:SEC Translocation Channels; C497347:SecM protein, E coli; D014157:Transcription Factors; C096222:tmRNA; D012343:RNA, Transfer; D000251:Adenosine Triphosphatases; D000081416:SecA Proteins