title: Subunit-selective proteasome activity profiling uncovers uncoupled proteasome subunit activities during bacterial infections.
abstract: The proteasome is a nuclear-cytoplasmic proteolytic complex involved in nearly all regulatory pathways in plant cells. The three different catalytic activities of the proteasome can have different functions, but tools to monitor and control these subunits selectively are not yet available in plant science. Here, we introduce subunit-selective inhibitors and dual-color fluorescent activity-based probes for studying two of the three active catalytic subunits of the plant proteasome. We validate these tools in two model plants and use this to study the proteasome during plant-microbe interactions. Our data reveal that Nicotiana benthamiana incorporates two different paralogs of each catalytic subunit into active proteasomes. Interestingly, both β1 and β5 activities are significantly increased upon infection with pathogenic Pseudomonas syringae pv. tomato DC3000 lacking hopQ1-1 [PtoDC3000(ΔhQ)] whilst the activity profile of the β1 subunit changes. Infection with wild-type PtoDC3000 causes proteasome activities that range from strongly induced β1 and β5 activities to strongly suppressed β5 activities, revealing that β1 and β5 activities can be uncoupled during bacterial infection. These selective probes and inhibitors are now available to the plant science community, and can be widely and easily applied to study the activity and role of the different catalytic subunits of the proteasome in different plant species.
authors.affiliation: The Plant Chemetics Laboratory, Max Planck Institute for Plant Breeding Research, Carl-von-Linné Weg 10, 50829, Cologne, Germany.|Botanical Institute and Cluster of Excellence on Plant Sciences, University of Cologne, 50674, Cologne, Germany.
authors.forename: Johana C
authors.initials: JC
authors.lastname: Misas-Villamil
authors.affiliation: The Plant Chemetics Laboratory, Max Planck Institute for Plant Breeding Research, Carl-von-Linné Weg 10, 50829, Cologne, Germany.
authors.forename: Aranka M
authors.initials: AM
authors.lastname: van der Burgh
authors.affiliation: The Plant Chemetics Laboratory, Department of Plant Sciences, University of Oxford, South Parks Lane, Oxford, OX1 3RB, UK.
authors.forename: Friederike
authors.initials: F
authors.lastname: Grosse-Holz
authors.affiliation: The Plant Chemetics Laboratory, Department of Plant Sciences, University of Oxford, South Parks Lane, Oxford, OX1 3RB, UK.
authors.forename: Marcel
authors.initials: M
authors.lastname: Bach-Pages
authors.affiliation: The Plant Chemetics Laboratory, Max Planck Institute for Plant Breeding Research, Carl-von-Linné Weg 10, 50829, Cologne, Germany.|Department of Plant Biology, University of Szeged, Szeged, Hungary.
authors.forename: Judit
authors.initials: J
authors.lastname: Kovács
authors.affiliation: Chemical Biology, Universität Duisburg-Essen, Zentrum für Medizinische Biotechnologie, Fakultät für Biologie, Universitätsstr. 2, 45117, Essen, Germany.
authors.forename: Farnusch
authors.initials: F
authors.lastname: Kaschani
authors.affiliation: The Plant Chemetics Laboratory, Max Planck Institute for Plant Breeding Research, Carl-von-Linné Weg 10, 50829, Cologne, Germany.
authors.forename: Sören
authors.initials: S
authors.lastname: Schilasky
authors.affiliation: The Plant Chemetics Laboratory, Max Planck Institute for Plant Breeding Research, Carl-von-Linné Weg 10, 50829, Cologne, Germany.
authors.forename: Asif E K
authors.initials: AE
authors.lastname: Emon
authors.affiliation: Gorlaeus Laboratories, Institute of Chemistry and Netherlands Proteomics Centre, 2333 CC, Leiden, The Netherlands.
authors.forename: Mark
authors.initials: M
authors.lastname: Ruben
authors.affiliation: Chemical Biology, Universität Duisburg-Essen, Zentrum für Medizinische Biotechnologie, Fakultät für Biologie, Universitätsstr. 2, 45117, Essen, Germany.
authors.forename: Markus
authors.initials: M
authors.lastname: Kaiser
authors.affiliation: Gorlaeus Laboratories, Institute of Chemistry and Netherlands Proteomics Centre, 2333 CC, Leiden, The Netherlands.
authors.forename: Hermen S
authors.initials: HS
authors.lastname: Overkleeft
authors.affiliation: The Plant Chemetics Laboratory, Max Planck Institute for Plant Breeding Research, Carl-von-Linné Weg 10, 50829, Cologne, Germany.|The Plant Chemetics Laboratory, Department of Plant Sciences, University of Oxford, South Parks Lane, Oxford, OX1 3RB, UK.
authors.forename: Renier A L
authors.initials: RA
authors.lastname: van der Hoorn
mesh_terms: D017360:Arabidopsis / Q000378:metabolism* / Q000382:microbiology*; D001424:Bacterial Infections / Q000378:metabolism*; D010935:Plant Diseases / Q000382:microbiology*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D044224:Pseudomonas syringae / Q000472:pathogenicity*; D014026:Nicotiana / Q000378:metabolism* / Q000382:microbiology*
chemical_list: D046988:Proteasome Endopeptidase Complex
keywords: 
  Arabidopsis thaliana
  ; 
  Nicotiana benthamiana
  ; activity-based protein profiling; catalytic subunit; core protease; proteasome manipulation; technical advance