Heat shock protein 90 and its co-chaperone protein phosphatase 5 interact with distinct regions of the tomato I-2 disease resistance protein.
Recent data suggest that plant disease resistance (R) proteins are present in multi-protein complexes. Tomato R protein I-2 confers resistance against the fungal pathogen Fusarium oxysporum. To identify components of the I-2 complex, we performed yeast two-hybrid screens using the I-2 leucine-rich repeat (LRR) domain as bait, and identified protein phosphatase 5 (PP5) as an I-2 interactor. Subsequent screens revealed two members of the cytosolic heat shock protein 90 (HSP90) family as interactors of PP5. By performing in vitro protein-protein interaction analysis using recombinant proteins, we were able to show a direct interaction between I-2 and PP5, and between I-2 and HSP90. The N-terminal part of the LRR domain was found to interact with HSP90, whereas the C-terminal part bound to PP5. The specific binding of HSP90 to the N-terminal region of the I-2 LRR domain was confirmed by co-purifying HSP90 from tomato lysate using recombinant proteins. Similarly, the interaction between PP5 and HSP90 was established. To investigate the role of PP5 and HSP90 for I-2 function, virus-induced gene silencing was performed in Nicotiana benthamiana. Silencing of HSP90 but not of PP5 completely blocked cell death triggered by I-2, showing that HSP90 is required for I-2 function. Together these data suggest that R proteins require, like steroid hormone receptors in animal systems, an HSP90/PP5 complex for their folding and functioning.
de la Fuente van Bentem
Sergio
S
Plant Pathology, Swammerdam Institute for Life Sciences, University of Amsterdam, Kruislaan 318, 1098 SM Amsterdam, The Netherlands.
Vossen
Jack H
JH
de Vries
Klaas Jan
KJ
van Wees
Saskia
S
Tameling
Wladimir I L
WI
Dekker
Henk L
HL
de Koster
Chris G
CG
Haring
Michel A
MA
Takken
Frank L W
FL
Cornelissen
Ben J C
BJ
D017360:Arabidopsis / Q000378:metabolism; D016923:Cell Death; D005670:Fusarium; D018506:Gene Expression Regulation, Plant; D020868:Gene Silencing; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism*; D007113:Immunity, Innate / Q000235:genetics; D018551:Solanum lycopersicum / Q000378:metabolism*; D018832:Molecular Chaperones / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism*; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D010935:Plant Diseases; D018515:Plant Leaves / Q000382:microbiology / Q000502:physiology; D010940:Plant Proteins / Q000378:metabolism*; D010942:Plant Viruses; D014026:Nicotiana / Q000378:metabolism; D020798:Two-Hybrid System Techniques
D018841:HSP90 Heat-Shock Proteins; D018832:Molecular Chaperones; D009687:Nuclear Proteins; D010940:Plant Proteins; D010749:Phosphoprotein Phosphatases; C089641:protein phosphatase 5