Production of Biologically Active Cecropin A Peptide in Rice Seed Oil Bodies

Montesinos, Laura; Bundó, Mireia; Izquierdo, Esther; Campo, Sonia; Badosa, Esther; Rossignol, Michel; Montesinos, E.; Segundo, Blanca San; Coca, María

doi:10.1371/journal.pone.0146919

Cited by 28 publications

(33 citation statements)

References 67 publications

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“…Despite a rise (130%) in rice production from 1966 to 2000 (Khush, ), output must accelerate to 852 million tons by 2035 to keep pace with the growing global population (Khush, ). To this end, genes that can improve grain yield and size of this staple crop have been intensively sought (Song et al ., ; Shomura et al ., ; Mao et al ., ; Li et al ., ; Wang et al ., ,b; Montesinos et al ., ; Liu et al ., ).…”

Section: Introductionmentioning

confidence: 99%

The overexpression of rice ACYL‐CoA‐BINDING PROTEIN2 increases grain size and bran oil content in transgenic rice

et al. 2019

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As Oryza sativa (rice) seeds represent food for over three billion people worldwide, the identification of genes that enhance grain size and composition is much desired. Past reports have indicated that Arabidopsis thaliana acyl-CoA-binding proteins (ACBPs) are important in seed development but did not affect seed size. Herein, rice OsACBP2 was demonstrated not only to play a role in seed development and germination, but also to influence grain size. OsACBP2 mRNA accumulated in embryos and endosperm of germinating seeds in qRT-PCR analysis, while b-glucuronidase (GUS) assays on OsACBP2pro::GUS rice transformants showed GUS expression in embryos, as well as the scutellum and aleurone layer of germinating seeds. Deletion analysis of the OsACBP2 5'-flanking region revealed five copies of the seed cis-element, Skn-I-like motif (À1486/À1482, À956/À952, À939/À935, À826/À822, and À766/À762), and the removal of any adversely affected expression in seeds, thereby providing a molecular basis for OsACBP2 expression in seeds. When OsACBP2 function was investigated using osacbp2 mutants and transgenic rice overexpressing OsACBP2 (OsACBP2-OE), osacbp2 was retarded in germination, while OsACBP2-OEs performed better than the wild-type and vector-transformed controls, in germination, seedling growth, grain size and grain weight. Transmission electron microscopy of OsACBP2-OE mature seeds revealed an accumulation of oil bodies in the scutellum cells, while confocal laser scanning microscopy indicated oil accumulation in OsACBP2-OE aleurone tissues. Correspondingly, OsACBP2-OE seeds showed gain in triacylglycerols and long-chain fatty acids over the vector-transformed control. As dietary rice bran contains beneficial bioactive components, OsACBP2 appears to be a promising candidate for enriching seed nutritional value.

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Section: Introductionmentioning

confidence: 99%

The overexpression of rice ACYL‐CoA‐BINDING PROTEIN2 increases grain size and bran oil content in transgenic rice

et al. 2019

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“…We found that rice-derived peptides increased intracellular glutathione levels. Rice is widely consumed around the world and rice-derived peptides are reported to have various biological effects such as anti-microbial activity, (8,9) anti-oxidative activity (10,11) and dipeptidyl peptidase-IV-inhibitory activity. (12,13) Compounds that have so far been reported to increase intracellular glutathione levels include silymarin, (14) cyanohydroxybutene, (15) α-naphthoflavone, (16) apocynin, (17) epalrestat, (18,19) ribose-cysteine, (20) β-carotene, (21) noradrenaline, (22) and lactacystin.…”

Section: Introductionmentioning

confidence: 99%

Hepatoprotective effects of rice-derived peptides against acetaminophen-induced damage in mice

Kawakami

Moritani

Uraji³

et al. 2017

J. Clin. Biochem. Nutr.

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Glutathione, the most abundant intracellular antioxidant, protects cells against reactive oxygen species induced oxidative stress and regulates intracellular redox status. We found that rice peptides increased intracellular glutathione levels in human hepatoblastoma HepG2 cells. Acetaminophen is a commonly used analgesic. However, an overdose of acetaminophen causes severe hepatotoxicity via depletion of hepatic glutathione. Here, we investigated the protective effects of rice peptides on acetaminophen-induced hepatotoxicity in mice. ICR mice were orally administered rice peptides (0, 100 or 500 mg/kg) for seven days, followed by the induction of hepatotoxicity via intraperitoneal injection of acetaminophen (700 mg/kg). Pretreatment with rice peptides significantly prevented increases in serum alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase levels and protected against hepatic glutathione depletion. The expression of γ-glutamylcysteine synthetase, a key regulatory enzyme in the synthesis of glutathione, was decreased by treatment with acetaminophen, albeit rice peptides treatment recovered its expression compared to that achieved treatment with acetaminophen. In addition, histopathological evaluation of the livers also revealed that rice peptides prevented acetaminophen-induced centrilobular necrosis. These results suggest that rice peptides increased intracellular glutathione levels and could protect against acetaminophen-induced hepatotoxicity in mice.

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“…We designed two different constructs to target this protein to two different subcellular compartments, namely apoplast and endoplasmic reticulum (ER). Compartmentalizing antimicrobial peptides in plant cells is known to favor their accumulation by protecting them from host proteases and avoiding their toxic effects on the host cells (Bundó et al ., ; Coca et al ., ; Montesinos et al ., ). To target AfpB to the apoplast or ER, we added the signal peptide of the tobacco AP24 protein () to the amino terminus of the mature AfpB protein (Figure a).…”

Section: Resultsmentioning

confidence: 97%

“…However, the yields reported when using these systems were much lower than the AFP yield achieved in N. benthamiana using the TMV‐derived system reported here. While our estimated yield of AfpB was approximately 250 μg per gram of fresh N. benthamiana leaf tissue, the reported yields for Retrocyclin‐101 and Protegrin‐1 were ~5 and 8 μg/g of tobacco respectively (Lee et al ., ), those for Cecropin A were ~40 μg/g of rice seeds (Montesinos et al ., ), those for LL37 were ~0.55 μg/g of barley seeds (Holásková et al ., ), and those for Lactoferrin chimera were ~4.8 μg/g of tobacco hairy roots (Chahardoli et al ., ). In addition, all of these plant‐based production systems are based on stable transformation, which makes the process more complex and time consuming.…”

Section: Discussionmentioning

confidence: 98%

“…While challenging due to the physicochemical properties and toxicity of many of these peptides, some of these platforms successfully produce AMPs. These platforms include leaf chloroplasts (Lee et al ., ), rice seeds (Bundó et al ., ; Montesinos et al ., , ), barley seeds (Holásková et al ., ) and hairy roots (Chahardoli et al ., ). However, the yields reported when using these systems were much lower than the AFP yield achieved in N. benthamiana using the TMV‐derived system reported here.…”

Section: Discussionmentioning

confidence: 99%

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Efficient production of antifungal proteins in plants using a new transient expression vector derived from tobacco mosaic virus

Shi

Cordero

Garrigues

et al. 2018

Plant Biotechnology Journal

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Summary Fungi that infect plants, animals or humans pose a serious threat to human health and food security. Antifungal proteins ( AFP s) secreted by filamentous fungi are promising biomolecules that could be used to develop new antifungal therapies in medicine and agriculture. They are small highly stable proteins with specific potent activity against fungal pathogens. However, their exploitation requires efficient, sustainable and safe production systems. Here, we report the development of an easy‐to‐use, open access viral vector based on Tobacco mosaic virus ( TMV ). This new system allows the fast and efficient assembly of the open reading frames of interest in small intermediate entry plasmids using the Gibson reaction. The manipulated TMV fragments are then transferred to the infectious clone by a second Gibson assembly reaction. Recombinant proteins are produced by agroinoculating plant leaves with the resulting infectious clones. Using this simple viral vector, we have efficiently produced two different AFP s in Nicotiana benthamiana leaves, namely the Aspergillus giganteus AFP and the Penicillium digitatum AfpB. We obtained high protein yields by targeting these bioactive small proteins to the apoplastic space of plant cells. However, when AFP s were targeted to intracellular compartments, we observed toxic effects in the host plants and undetectable levels of protein. We also demonstrate that this production system renders AFP s fully active against target pathogens, and that crude plant extracellular fluids containing the AfpB can protect tomato plants from Botrytis cinerea infection, thus supporting the idea that plants are suitable biofactories to bring these antifungal proteins to the market.

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Production of Biologically Active Cecropin A Peptide in Rice Seed Oil Bodies

Cited by 28 publications

References 67 publications

The overexpression of rice ACYL‐CoA‐BINDING PROTEIN2 increases grain size and bran oil content in transgenic rice

The overexpression of rice ACYL‐CoA‐BINDING PROTEIN2 increases grain size and bran oil content in transgenic rice

Hepatoprotective effects of rice-derived peptides against acetaminophen-induced damage in mice

Efficient production of antifungal proteins in plants using a new transient expression vector derived from tobacco mosaic virus

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