Self-assembled poloxamer-legumin/vicilin nanoparticles for the nanoencapsulation and controlled release of folic acid.

Plant-based food proteins are a promising choice for the preparation of nanoparticles (NPs) due to their high digestibility, low cost, and ability to interact with various compounds and nutrients. Moreover, nanoencapsulation offers a potential solution for protecting nutrients during processing and enhancing their bioavailability. This study aimed to develop and evaluate nanoparticles (NPs) based on legumin/vicilin (LV) proteins extracted from fava beans, with the goal of encapsulating and delivering a model nutraceutical compound, folic acid (FA). Specifically, NPs were self-assembled from LV proteins extracted from commercially available frozen fava beans using a pH-coacervation method with poloxamer 188 (P188) and chemically cross-linked with glutaraldehyde. Microscopy and spectroscopy studies were carried out on the empty and FA-loaded NPs in order to evaluate the particle morphology, size, size distribution, composition, mechanism of formation, impact of FA loading and release behavior. In vitro studies with Caco-2 cells also confirmed that the empty and FA-loaded nanoparticles were non-toxic. Thus, the LV-NPs are good candidates as food additives for the delivery and stabilization of nutrients as well as in drug delivery for the controlled release of therapeutics.

Proximity-dependent biotinylation screening identifies NbHYPK as a novel interacting partner of ATG8 in plants.

BACKGROUND: Autophagy is a conserved, highly-regulated catabolic process that plays important roles in growth, development and innate immunity in plants. In this study, we compared the rate of autophagy induction in Nicotiana benthamiana plants infected with Tobacco mosaic virus or the TMV 24A + UPD mutant variant, which replicates at a faster rate and induces more severe symptoms. Using a BirA* tag and proximity-dependent biotin identification (BioID) analysis, we identified host proteins that interact with the core autophagy protein, ATG8 in TMV 24A + UPD infected plants. By combining the use of a fast replicating TMV mutant and an in vivo protein-protein screening technique, we were able to gain functional insight into the role of autophagy in a compatible virus-host interaction. RESULTS: Our study revealed an increased autophagic flux induced by TMV 24A + UPD, as compared to TMV in N. benthamiana. Analysis of the functional proteome associated with ATG8 revealed a total of 67 proteins, 16 of which are known to interact with ATG8 or its orthologs in mammalian and yeast systems. The interacting proteins were categorized into four functional groups: immune system process, response to ROS, sulphur amino acid metabolism and calcium signalling. Due to the presence of an ubiquitin-associated (UBA) domain, which is demonstrated to interact with ATG8, the Huntingtin-interacting protein K-like (HYPK) was selected for validation of the physical interaction and function. We used yeast two hybrid (Y2H), bimolecular fluorescence complementation (BiFC) and subcellular localization to validate the ATG8-HYPK interaction. Subsequent down-regulation of ATG8 by virus-induced gene silencing (VIGS) showed enhanced TMV symptoms, suggesting a protective role for autophagy during TMV 24A + UPD infection. CONCLUSION: This study presents the use of BioID as a suitable method for screening ATG8 interacting proteins in planta. We have identified many putative binding partners of ATG8 during TMV 24A + UPD infection in N. benthamiana plants. In addition, we have verified that NbHYPK is an interacting partner of ATG8. We infer that autophagy plays a protective role in TMV 24A + UPD infected plants.