Oversynthesis of a new Escherichia coli small RNA suppresses export toxicity of DsbA'-PhoA unfoldable periplasmic proteins.
In Escherichia coli, the DsbA'-PhoA hybrid proteins carrying an unfoldable DsbA' fragment can be targeted to the envelope, where they exert their toxicity. Hybrid proteins stick to the periplasmic face of the inner membrane and paralyze the export mechanism, becoming lethal if sufficiently overproduced and if not degraded by the DegP protease (A. Guigueno, P. Belin, and P. L. Boquet, J. Bacteriol. 179:3260-3269, 1997). We isolated a multicopy suppressor that restores viability to a degP strain without modifying the expression level of the toxic fusion. Suppression does not involve activation of the known envelope stress-combative pathways, the Cpx pathway and the sigma(E) regulon. Subclone analysis of the suppressor revealed a 195-bp DNA fragment that is responsible for toxicity suppression. The cloned gene, called uptR, is approximately 130 bp long (including the promoter and a transcription termination signal) and is transcribed into a small RNA (92 nucleotides). Using site-directed mutagenesis, we found that UptR RNA does not require translation for toxicity suppression. UptR-mediated action reduces the amount of membrane-bound toxic hybrid protein. UptR RNA is the first example of a small RNA implicated in extracytoplasmic toxicity suppression. It appears to offer a new way of suppressing toxicity, and its possible modes of action are discussed.
Guigueno
A
A
Département d'Ingénierie et d'Etudes des Protéines, Commissariat à l'Energie Atomique, 91191 Gif-sur-Yvette, France.
Dassa
J
J
Belin
P
P
Boquet
P L
PL
D000469:Alkaline Phosphatase / Q000378:metabolism / Q000633:toxicity; D001426:Bacterial Proteins / Q000378:metabolism / Q000633:toxicity*; D001483:Base Sequence; D003001:Cloning, Molecular; D004275:DNA, Ribosomal; D004926:Escherichia coli / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D008565:Membrane Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D019897:Periplasm / Q000378:metabolism*; D019704:Protein Disulfide-Isomerases / Q000378:metabolism / Q000633:toxicity; D017510:Protein Folding; D021381:Protein Transport / Q000235:genetics*; D012329:RNA, Bacterial / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism / Q000633:toxicity; D017422:Sequence Analysis, DNA; D012808:Sigma Factor / Q000378:metabolism; D013489:Suppression, Genetic; D014157:Transcription Factors / Q000378:metabolism
D001426:Bacterial Proteins; D004275:DNA, Ribosomal; D008565:Membrane Proteins; D012329:RNA, Bacterial; D011993:Recombinant Fusion Proteins; D012808:Sigma Factor; D014157:Transcription Factors; C063202:sporulation-specific sigma factors; D000469:Alkaline Phosphatase; D019704:Protein Disulfide-Isomerases