Intracellular Delivery of Proteins via Fusion Peptides in Intact Plants.
In current plant biotechnology, the introduction of exogenous DNA encoding desired traits is the most common approach used to modify plants. However, general plant transformation methods can cause random integration of exogenous DNA into the plant genome. To avoid these events, alternative methods, such as a direct protein delivery system, are needed to modify the plant. Although there have been reports of the delivery of proteins into cultured plant cells, there are currently no methods for the direct delivery of proteins into intact plants, owing to their hierarchical structures. Here, we demonstrate the efficient fusion-peptide-based delivery of proteins into intact Arabidopsis thaliana. Bovine serum albumin (BSA, 66 kDa) was selected as a model protein to optimize conditions for delivery into the cytosol. The general applicability of our method to large protein cargo was also demonstrated by the delivery of alcohol dehydrogenase (ADH, 150 kDa) into the cytosol. The compatibility of the fusion peptide system with the delivery of proteins to specific cellular organelles was also demonstrated using the fluorescent protein Citrine (27 kDa) conjugated to either a nuclear localization signal (NLS) or a peroxisomal targeting signal (PTS). In conclusion, our designed fusion peptide system can deliver proteins with a wide range of molecular weights (27 to 150 kDa) into the cells of intact A. thaliana without interfering with the organelle-targeting peptide conjugated to the protein. We expect that this efficient protein delivery system will be a powerful tool in plant biotechnology.
Enzyme Research Team, Biomass Engineering Research Division, RIKEN Center for Sustainable Resource Science, Wako, Saitama, Japan.|School of Biological Sciences, Universiti Sains Malaysia, Penang, Malaysia.
Kiaw Kiaw
KK
Ng
Enzyme Research Team, Biomass Engineering Research Division, RIKEN Center for Sustainable Resource Science, Wako, Saitama, Japan.|Laboratory for Biomolecular Structure and Dynamics, RIKEN Quantitative Biology Center (QBiC), Yokohama, Kanagawa, Japan.
Yoko
Y
Motoda
Laboratory for Biomolecular Structure and Dynamics, RIKEN Quantitative Biology Center (QBiC), Yokohama, Kanagawa, Japan.
Satoru
S
Watanabe
School of Biological Sciences, Universiti Sains Malaysia, Penang, Malaysia.
Ahmad
A
Sofiman Othman
Laboratory for Biomolecular Structure and Dynamics, RIKEN Quantitative Biology Center (QBiC), Yokohama, Kanagawa, Japan.|Department of Computational Intelligence and Systems Science, Interdisciplinary Graduate School of Science and Engineering, Tokyo Institute of Technology, Yokohama, Kanagawa, Japan.
Takanori
T
Kigawa
Center for Bioscience Research and Education, Utsunomiya University, Utsunomiya, Tochigi, Japan.
Yutaka
Y
Kodama
Enzyme Research Team, Biomass Engineering Research Division, RIKEN Center for Sustainable Resource Science, Wako, Saitama, Japan.
Keiji
K
Numata
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000378:metabolism*; D008970:Molecular Weight; D015388:Organelles / Q000378:metabolism; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism*; D012710:Serum Albumin, Bovine / Q000008:administration & dosage*
D010455:Peptides; D011993:Recombinant Fusion Proteins; D012710:Serum Albumin, Bovine