Determining the Size Exclusion for Nanoparticles in Citrus Leaves

Implementation of nanotechnology in agriculture is intimately dependent on the capacity of nanoparticles (NPs) to move within the plant body and reach the targeted cells. Although the fibrillar nature of the plant cell wall permits the movement of molecules through its porous matrix (apoplast), the movement of particles through the aqueous apoplastic milieu has its size limitations given the tightly knitted cellulose/hemicellulose fiber structure. In the present study, we used fluorescent NPs of different composition and sizes, and followed their movement into citrus leaves by fluorescent microscopy. Our results indicate that in citrus leaves, the size exclusion limit for NPs is of ≈5.4 nm. This conclusion was based on the capacity of PAMAM dendrimers G-4 and G-5 (4.5 and 5.4 nm, respectively) to move through the cell wall and into the phloem, but failure of similar PAMAM dendrimers G-6 (6.7 nm) to move through the apoplast. Dendrimer NPs 5.4 nm and smaller were observed to penetrate the leaf tissue, and then taken up and mobilized by the phloem elements. The current study provides evidence on the size limit for NPs use in agriculture.

Identification of a developmental transition in plasmodesmatal function during embryogenesis in Arabidopsis thaliana

Plasmodesmata provide routes for communication and nutrient transfer between plant cells by interconnecting the cytoplasm of adjacent cells. A simple fluorescent tracer loading assay was developed to monitor patterns of cell-to-cell transport via plasmodesmata specifically during embryogenesis. A developmental transition in plasmodesmatal size exclusion limit was found to occur at the torpedo stage of embryogenesis in Arabidopsis; at this time, plasmodesmata are down-regulated, allowing transport of small (approx. 0.5 kDa) but not large (approx. 10 kDa) tracers. This assay system was used to screen for embryo-defective mutants, designated increased size exclusion limit of plasmodesmata(ise), that maintain dilated plasmodesmata at the torpedo stage. The morphology of ise1 and ise2 mutants discussed here resembled that of the wild-type during embryo development, although the rate of their embryogenesis was slower. The ISE1 gene was mapped to position 13 cM on chromosome I using PCR-based biallelic markers. ise2 was found to be allelic to the previously characterized mutant emb25 which maps to position 100 cM on chromosome I. The results presented have implications for intercellular signaling pathways that regulate embryonic development, and furthermore represent the first attempt to screen directly for mutants of Arabidopsis with altered size exclusion limit of plasmodesmata.

Structure and function of plasmodesmata

Little is known about the molecular architecture of plasmodesmata. Electron micrographs of plasmodesmata, although showing remarkably consistent features, do not reveal the dynamic nature of plasmodesmata. In addition, the generally accepted size exclusion limit of plasmodesmata of 1 kDa as determined by the microinjection of fluorescent probes, is not an accurate measure of the size of molecules that can move from cell to cell. In some cases, plasmodesmata allow the non-selective movement of proteins up to 50 kDa, whereas others allow the selective transport of proteins such as transcription factors. Insights to the molecular architecture of plasmodesmata and the mechanism of cell-to-cell transport can be gained by the examination of the type of molecules and conditions under which selective and passive movement of molecules occurs.