Sugar Promotes Vegetative Phase Change in \u3cem\u3eArabidopsis thaliana\u3c/em\u3e by Repressing the Expression of \u3cem\u3eMIR156A\u3c/em\u3e and \u3cem\u3eMIR156C\u3c/em\u3e

Nutrients shape the growth, maturation, and aging of plants and animals. In plants, the juvenile to adult transition (vegetative phase change) is initiated by a decrease in miR156. In Arabidopsis, we found that exogenous sugar decreased the abundance of miR156, whereas reduced photosynthesis increased the level of this miRNA. This effect was correlated with a change in the timing of vegetative phase change, and was primarily attributable to a change in the expression of two genes, MIR156A and MIR156C, which were found to play dominant roles in this transition. The glucose-induced repression of miR156 was dependent on the signaling activity of HEXOKINASE1. We also show that the defoliation-induced increase in miR156 levels can be suppressed by exogenous glucose. These results provide a molecular link between nutrient availability and developmental timing in plants, and suggest that sugar is a component of the leaf signal that mediates vegetative phase chang

Optimized protocols for protoplast isolation, transfection, and regeneration in the Solanum genus for the CRISPR/Cas-mediated transgene-free genome editing

Abstract The Solanaceae family includes the largest flowering crops such as tomatoes, potatoes, and eggplants. Consumer demand has led to massive development of plants in the Solanum genus, and many different Solanum varieties are now available on the market. The recent advances in Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/CRISPR-associated protein 9 (Cas9)-based genome editing have allowed laboratories and smaller crop production companies to utilize the technology in various crops. The traditional transformation method in crops involves the use of Agrobacterium, which is considered the most efficient method for introducing exogenous genetic materials in target plants. The Agrobacterium-mediated transformation method has been also established in the Solanaceae family, enabling CRISPR/Cas-based genome editing in crops like tomatoes, potatoes, and eggplants. However, the Agrobacterium-mediated approach inevitably accompanies the insertion of exogenous DNA into the plant genome and often causes the formation of chimera that require further propagation steps. Alternatively, the CRISPR/Cas components can be introduced into protoplasts in the form of DNA for transient expression or a mixture of protein and RNA to avoid genomic insertion of foreign materials. The protoplast transformation approach involves processes including protoplast preparation, transfection, and regeneration, which require a comprehensive understanding and greater technical mastery of the tissue culture phase. Here we highlight the current research advances in protoplast transformation and discuss how to optimize the procedures of protoplast isolation, transfection, and regeneration for efficient and reproducible CRISPR/Cas-based genome editing in the genus Solanum

Fluorescence Reports Intact Quantum Dot Uptake into Roots and Translocation to Leaves of <i>Arabidopsis thaliana</i> and Subsequent Ingestion by Insect Herbivores

We explored the impact of quantum dot (QD) coat characteristics on NP stability, uptake, and translocation in <i>Arabidopsis thaliana</i>, and subsequent transfer to primary consumers, <i>Trichoplusia ni</i> (<i>T. ni</i>). <i>Arabidopsis</i> was exposed to CdSe/CdZnS QDs with three different coatings: Poly­(acrylic acid-ethylene glycol) (PAA-EG), polyethylenimine (PEI) and poly­(maleic anhydride-<i>alt</i>-1-octadecene)–poly­(ethylene glycol) (PMAO–PEG), which are anionic, cationic, and relatively neutral, respectively. PAA-EG-coated QDs were relatively stable and taken up from a hydroponic medium through both <i>Arabidopsis</i> leaf petioles and roots, without apparent aggregation, and showed generally uniform distribution in leaves. In contrast, PEI- and PMAO–PEG-coated QDs displayed destabilization in the hydroponic medium, and generated particulate fluorescence plant tissues, suggesting aggregation. PAA-EG QDs moved faster than PEI QDs through leaf petioles; however, 8-fold more cadmium accumulated in PEI QD-treated leaves than in those exposed to PAA-EG QDs, possibly due to PEI QD dissolution and direct metal uptake. <i>T. ni</i> caterpillars that fed on <i>Arabidopsis</i> exposed to QDs had reduced performance, and QD fluorescence was detected in both <i>T. ni</i> bodies and frass, demonstrating trophic transfer of intact QDs from plants to insects. Overall, this paper demonstrates that QD coat properties influence plant nanoparticle uptake and translocation and can impact transfer to herbivores

Characterization of and isolation methods for plant leaf nanovesicles and small extracellular vesicles

Mammalian small extracellular vesicles (sEVs) can deliver diverse molecules to target cells. However, they are difficult to obtain in large quantities and can activate host immune responses. Plant-derived vesicles may help to overcome these challenges. We optimized isolation methods for two types of plant vesicles, nanovesicles from disrupted leaf and sEVs from the extracellular apoplastic space of Arabidopsis thaliana. Both preparations yielded intact vesicles of uniform size, and a mean membrane charge of approximately -25 mV. We also demonstrated applicability of these preparative methods using Brassicaceae vegetables. Proteomic analysis of a subset of vesicles with a density of 1.1-1.19 g mL(-1) sheds light on the likely cellular origin and complexity of the vesicles. Both leaf nanovesicles and sEVs were taken up by cancer cells, with sEVs showing an approximately three-fold higher efficiency compared to leaf nanovesicles. These results support the potential of plant-derived vesicles as vehicles for therapeutic delivery. (c) 2020 Elsevier Inc. All rights reserved