Leaf proteome rebalancing in Nicotiana benthamiana for upstream enrichment of a transiently expressed recombinant protein.
A key factor influencing the yield of biopharmaceuticals in plants is the ratio of recombinant to host proteins in crude extracts. Postextraction procedures have been devised to enrich recombinant proteins before purification. Here, we assessed the potential of methyl jasmonate (MeJA) as a generic trigger of recombinant protein enrichment in Nicotiana benthamiana leaves before harvesting. Previous studies have reported a significant rebalancing of the leaf proteome via the jasmonate signalling pathway, associated with ribulose 1,5-bisphosphate carboxylase oxygenase (RuBisCO) depletion and the up-regulation of stress-related proteins. As expected, leaf proteome alterations were observed 7 days post-MeJA treatment, associated with lowered RuBisCO pools and the induction of stress-inducible proteins such as protease inhibitors, thionins and chitinases. Leaf infiltration with the Agrobacterium tumefaciens bacterial vector 24 h post-MeJA treatment induced a strong accumulation of pathogenesis-related proteins after 6 days, along with a near-complete reversal of MeJA-mediated stress protein up-regulation. RuBisCO pools were partly restored upon infiltration, but most of the depletion effect observed in noninfiltrated plants was maintained over six more days, to give crude protein samples with 50% less RuBisCO than untreated tissue. These changes were associated with net levels reaching 425 μg/g leaf tissue for the blood-typing monoclonal antibody C5-1 expressed in MeJA-treated leaves, compared to less than 200 μg/g in untreated leaves. Our data confirm overall the ability of MeJA to trigger RuBisCO depletion and recombinant protein enrichment in N. benthamiana leaves, estimated here for C5-1 at more than 2-fold relative to host proteins.
Centre de recherche et d'innovation sur les végétaux, Pavillon Envirotron, Université Laval, Québec, QC, Canada.
Stéphanie
S
Robert
Centre de recherche et d'innovation sur les végétaux, Pavillon Envirotron, Université Laval, Québec, QC, Canada.
Marie-Claire
MC
Goulet
Medicago Inc., Québec, QC, Canada.
Marc-André
MA
D'Aoust
Centre de recherche et d'innovation sur les végétaux, Pavillon Envirotron, Université Laval, Québec, QC, Canada.|The University of Queensland, Australian Institute for Bioengineering and Nanotechnology, St Lucia, QLD, Australia.
Frank
http://orcid.org/0000-0001-8152-3820
F
Sainsbury
Centre de recherche et d'innovation sur les végétaux, Pavillon Envirotron, Université Laval, Québec, QC, Canada.
Dominique
D
Michaud
D000085:Acetates / Q000494:pharmacology; D016960:Agrobacterium tumefaciens / Q000187:drug effects; D000906:Antibodies / Q000378:metabolism; D003517:Cyclopentanes / Q000494:pharmacology; D054883:Oxylipins / Q000494:pharmacology; D010447:Peptide Hydrolases / Q000378:metabolism; D018515:Plant Leaves / Q000187:drug effects / Q000378:metabolism*; D030821:Plants, Genetically Modified; D020543:Proteome / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis*; D014026:Nicotiana / Q000187:drug effects / Q000235:genetics*; D014162:Transfection; D015854:Up-Regulation / Q000187:drug effects
D000085:Acetates; D000906:Antibodies; D003517:Cyclopentanes; D054883:Oxylipins; D020543:Proteome; D011994:Recombinant Proteins; C072239:methyl jasmonate; D010447:Peptide Hydrolases
Nicotiana benthamiana; agroinfiltration; leaf proteome; methyl jasmonate; molecular farming; recombinant protein yield; transient expression