The PscE-PscF-PscG complex controls type III secretion needle biogenesis in Pseudomonas aeruginosa.
Type III secretion (T3S) systems play key roles in pathogenicity of many Gram-negative bacteria and are employed to inject toxins directly into the cytoplasm of target cells. They are composed of over 20 different proteins that associate into a basal structure that traverses both inner and outer bacterial membranes and a hollow, needle-like structure through which toxins travel. The PscF protein is the main component of the Pseudomonas aeruginosa T3S needle. Here we demonstrate that PscF, when purified on its own, is able to form needle-like fibers of 8 nm in width and >1 microm in length. In addition, we demonstrate for the first time that the T3S needle subunit requires two cytoplasmic partners, PscE and PscG, in P. aeruginosa, which trap PscF in a ternary, 1:1:1 complex, thus blocking it in a monomeric state. Knock-out mutants deficient in PscE and PscG are non-cytotoxic, lack PscF, and are unable to export PscF encoded extrachromosomally. Temperature-scanning circular dichroism measurements show that the PscE-PscF-PscG complex is thermally stable and displays a cooperative unfolding/refolding pattern. Thus, PscE and PscG prevent PscF from polymerizing prematurely in the P. aeruginosa cytoplasm and keep it in a secretion prone conformation, strategies which may be shared by other pathogens that employ the T3S system for infection.
Quinaud
Manuelle
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Biochimie et Biophysique des Systèmes Intégrés, UMR 5092 CNRS/Commissariat à l'Energie Atomique (CEA)/Université Joseph Fourier (UJF), Département de Réponse et Dynamique Cellulaires, CEA Grenoble, 17 rue des Martyrs, 38054 Grenoble cedex 09, France.
Chabert
Jacqueline
J
Faudry
Eric
E
Neumann
Emmanuelle
E
Lemaire
David
D
Pastor
Alexandrine
A
Elsen
Sylvie
S
Dessen
Andréa
A
Attree
Ina
I
D000595:Amino Acid Sequence; D000942:Antigens, Bacterial / Q000737:chemistry; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry; D018407:Bacterial Physiological Phenomena; D001426:Bacterial Proteins / Q000737:chemistry; D015153:Blotting, Western; D002352:Carrier Proteins / Q000235:genetics / Q000502:physiology*; D002942:Circular Dichroism; D003593:Cytoplasm / Q000378:metabolism; D005822:Genetic Vectors; D015151:Immunoblotting; D036341:Intercellular Signaling Peptides and Proteins; D013058:Mass Spectrometry; D026901:Membrane Transport Proteins / Q000737:chemistry; D008854:Microscopy, Electron; D018832:Molecular Chaperones / Q000235:genetics / Q000502:physiology*; D008969:Molecular Sequence Data; D009154:Mutation; D011108:Polymers / Q000737:chemistry; D011487:Protein Conformation; D011550:Pseudomonas aeruginosa / Q000378:metabolism*; D013696:Temperature; D013997:Time Factors; D019076:Transgenes
D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D002352:Carrier Proteins; D036341:Intercellular Signaling Peptides and Proteins; D026901:Membrane Transport Proteins; D018832:Molecular Chaperones; D011108:Polymers; C506610:PscE protein, Pseudomonas aeruginosa; C506611:PscF protein, Pseudomonas aeruginosa; C506612:PscG protein, Pseudomonas aeruginosa