title: A nonphosphorylated 14-3-3 binding motif on exoenzyme S that is functional in vivo.
abstract: 14-3-3 proteins play an important role in a multitude of signalling pathways. The interactions between 14-3-3 and other signalling proteins, such as Raf and KSR (kinase suppressor of Ras), occur in a phospho-specific manner. Recently, a phosphorylation-independent interaction has been reported to occur between 14-3-3 and several proteins, for example 5-phosphatase, p75NTR-associated cell death executor (NADE) and the bacterial toxin Exoenzyme S (ExoS), an ADP-ribosyltransferase from Pseudomonas aeruginosa. In this study we have identified the amino acid residues on ExoS, which are responsible for its specific interaction with 14-3-3. Furthermore, we show that a peptide derived from ExoS, containing the 14-3-3 interaction site, effectively competes out the interaction between ExoS and 14-3-3. In addition, competition with this peptide blocks ExoS modification of Ras in our Ras modification assay. We show that the ExoS protein interacts with all isoforms of the 14-3-3 family tested. Moreover, in vivo an ExoS protein lacking the 14-3-3 binding site has a reduced capacity to ADP ribosylate cytoplasmic proteins, e.g. Ras, and shows a reduced capacity to change the morphology of infected cells.
authors.affiliation: Department of Medical Biosciences/Pathology, Umeå University, Sweden.
authors.forename: Maria L
authors.initials: ML
authors.lastname: Henriksson
authors.forename: Matthew S
authors.initials: MS
authors.lastname: Francis
authors.forename: Alex
authors.initials: A
authors.lastname: Peden
authors.forename: Margareta
authors.initials: M
authors.lastname: Aili
authors.forename: Kristina
authors.initials: K
authors.lastname: Stefansson
authors.forename: Ruth
authors.initials: R
authors.lastname: Palmer
authors.forename: Alastair
authors.initials: A
authors.lastname: Aitken
authors.forename: Bengt
authors.initials: B
authors.lastname: Hallberg
mesh_terms: D048948:14-3-3 Proteins; D036002:ADP Ribose Transferases / Q000235:genetics / Q000378:metabolism*; D000244:Adenosine Diphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D001427:Bacterial Toxins / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D001667:Binding, Competitive; D006367:HeLa Cells / Q000166:cytology / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism; D010766:Phosphorylation; D020033:Protein Isoforms; D017384:Sequence Deletion; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism*; D018631:ras Proteins / Q000378:metabolism
chemical_list: D048948:14-3-3 Proteins; D001427:Bacterial Toxins; D010446:Peptide Fragments; D020033:Protein Isoforms; D000244:Adenosine Diphosphate; D014446:Tyrosine 3-Monooxygenase; D036002:ADP Ribose Transferases; C444820:exoenzyme S; D018631:ras Proteins