Oral Presentation International Plant Molecular Biology Conference 2024

Protein-coated layered double hydroxide (LDH) nanosheets for delivery of transgenes and synthetic mRNA into intact plants (107906)

Jiaxi Yong 1 2 , Wang Xu 2 , Miaomiao Wu 1 , Run Zhang 1 , Christopher W.G. Mann 2 , Guoquan Liu 3 , Christopher A. Brosnan 3 , Neena Mitter 3 , Bernard J Carroll 2 , Zhi Ping Xu 1 4
  1. Australian Institute for Bioengineering and Nanotechnology, The University of Queensland, Brisbane, QLD, Australia
  2. School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, QLD, Australia
  3. Queensland Alliance for Agriculture and Food Innovation, The University of Queensland, Brisbane, QLD, Australia
  4. Institute of Biomedical Health Technology and Engineering and Institute of Systems and Physical Biology, Shenzhen Bay Laboratory, Shenzhen, China

We have designed protein-coated nanoparticles that can effectively traverse the plant cell wall, and can be taken up and translocate via an active transport mechanism in plants. Using commercially available proteins, we discovered that lysozyme coating of the nanosheets greatly enhances their uptake and translocation through the roots.

The lysozyme-coated nanosheets are actively taken up into the root tip and elongation zone by endocytosis and translocate through the plant via a membrane trafficking pathway. We initially hypothesised that coating of the LDH nanosheets with any protein would enhance uptake via endocytosis. However, we found that a Bovine Serum Albumin (BSA) coating had the opposite effect and decreased nanoparticle uptake. Uncoated and lysozyme-coated nanosheets are positively charged (39 and 20 mV, respectively), whereas BSA-coated nanosheets have a negative surface charge (-18 mV), which most likely contributes to the differences in nanosheet uptake. Lysozyme is an antimicrobial enzyme in animals that breaks the glycosidic linkages in bacterial cell walls, but it also possesses a much weaker capacity to break β-1,4-glycosidic linkages in cellulose. Our data indicate that lysozyme coating greatly enhances nanosheet uptake by i) loosening the cell wall, and ii) stimulating expression of membrane trafficking genes. We have used our lysozyme-coated nanosheets to not only effectively

deliver transgenes but also synthetic mRNA into dicot and monocot plant species for the first time. The overarching aim of this project is to develop a non-GM platform for all plant species based on protein-coated nanoparticle delivery of nucleic acids into whole plants.