title: The E. coli effector protein NleF is a caspase inhibitor.
abstract: Enterohemorrhagic and enteropathogenic E. coli (EHEC and EPEC) can cause severe and potentially life-threatening infections. Their pathogenicity is mediated by at least 40 effector proteins which they inject into their host cells by a type-III secretion system leading to the subversion of several cellular pathways. However, the molecular function of several effectors remains unknown, even though they contribute to virulence. Here we show that one of them, NleF, binds to caspase-4, -8, and -9 in yeast two-hybrid, LUMIER, and direct interaction assays. NleF inhibits the catalytic activity of the caspases in vitro and in cell lysate and prevents apoptosis in HeLa and Caco-2 cells. We have solved the crystal structure of the caspase-9/NleF complex which shows that NleF uses a novel mode of caspase inhibition, involving the insertion of the carboxy-terminus of NleF into the active site of the protease. In conformance with our structural model, mutagenized NleF with truncated or elongated carboxy-termini revealed a complete loss in caspase binding and apoptosis inhibition. Evasion of apoptosis helps pathogenic E. coli and other pathogens to take over the host cell by counteracting the cell's ability to self-destruct upon infection. Recently, two other effector proteins, namely NleD and NleH, were shown to interfere with apoptosis. Even though NleF is not the only effector protein capable of apoptosis inhibition, direct inhibition of caspases by bacterial effectors has not been reported to date. Also unique so far is its mode of inhibition that resembles the one obtained for synthetic peptide-type inhibitors and as such deviates substantially from previously reported caspase-9 inhibitors such as the BIR3 domain of XIAP.
authors.affiliation: Genomics and Proteomics Core Facilities, German Cancer Research Center, Heidelberg, Germany.
authors.forename: Sonja
authors.initials: S
authors.lastname: Blasche
authors.forename: Mario
authors.initials: M
authors.lastname: Mörtl
authors.forename: Holger
authors.initials: H
authors.lastname: Steuber
authors.forename: Gabriella
authors.initials: G
authors.lastname: Siszler
authors.forename: Shahista
authors.initials: S
authors.lastname: Nisa
authors.forename: Frank
authors.initials: F
authors.lastname: Schwarz
authors.forename: Inna
authors.initials: I
authors.lastname: Lavrik
authors.forename: Thomas M A
authors.initials: TM
authors.lastname: Gronewold
authors.forename: Klaus
authors.initials: K
authors.lastname: Maskos
authors.forename: Michael S
authors.initials: MS
authors.lastname: Donnenberg
authors.forename: Dirk
authors.initials: D
authors.lastname: Ullmann
authors.forename: Peter
authors.initials: P
authors.lastname: Uetz
authors.forename: Manfred
authors.initials: M
authors.lastname: Kögl
mesh_terms: D061945:Caspase Inhibitors / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D020169:Caspases / Q000737:chemistry / Q000378:metabolism*; D002460:Cell Line; D019453:Escherichia coli O157 / Q000502:physiology; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D008958:Models, Molecular; D011487:Protein Conformation; D015203:Reproducibility of Results; D037521:Virulence Factors / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology
chemical_list: D061945:Caspase Inhibitors; D029968:Escherichia coli Proteins; C583422:NleF protein, E coli; D037521:Virulence Factors; D020169:Caspases