Production of BP178, a derivative of the synthetic antibacterial peptide BP100, in the rice seed endosperm

Abstract

BackgroundBP178 peptide is a synthetic BP100-magainin derivative possessing strong inhibitory activity against plant pathogenic bacteria, offering a great potential for future applications in plant protection and other fields. Here we report the production and recovery of a bioactive BP178 peptide using rice seeds as biofactories.ResultsA synthetic gene encoding the BP178 peptide was prepared and introduced in rice plants. The gene was efficiently expressed in transgenic rice under the control of an endosperm-specific promoter. Among the three endosperm-specific rice promoters (Glutelin B1, Glutelin B4 or Globulin 1), best results were obtained when using the Globulin 1 promoter. The BP178 peptide accumulated in the seed endosperm and was easily recovered from rice seeds using a simple procedure with a yield of 21 μg/g. The transgene was stably inherited for at least three generations, and peptide accumulation remained stable during long term storage of transgenic seeds. The purified peptide showed in vitro activity against the bacterial plant pathogen Dickeya sp., the causal agent of the dark brown sheath rot of rice. Seedlings of transgenic events showed enhanced resistance to the fungal pathogen Fusarium verticillioides, supporting that the in planta produced peptide was biologically active.ConclusionsThe strategy developed in this work for the sustainable production of BP178 peptide using rice seeds as biofactories represents a promising system for future production of peptides for plant protection and possibly in other fields.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-017-1011-9) contains supplementary material, which is available to authorized users.

Screening and identification of BP100 peptide conjugates active against Xylella fastidiosa using a viability-qPCR method

“…Within our search for new AMPs to control plant diseases, we reported peptide conjugates incorporating units of the lead peptide BP100 and fragments of cecropin A, magainin II or melittin, which were specifically designed to be expressed in plants [17,18]. In fact, the peptide conjugate BP178 was successfully expressed in rice endosperm, showing resistance against some plant pathogens [19]. This demonstrates the availability of these peptides for being produced by the plant itself, which could overcome the difficulties in accessing the vascular location of Xf observed by other treatment strategies.…”

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

“…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 lg per gram of fresh N. benthamiana leaf tissue, the reported yields for Retrocyclin-101 and Protegrin-1 were~5 and 8 lg/g of tobacco respectively (Lee et al, 2011), those for Cecropin A were~40 lg/ g of rice seeds (Montesinos et al, 2016), those for LL37 werẽ 0.55 lg/g of barley seeds (Hol askov a et al, 2018), and those for Lactoferrin chimera were~4.8 lg/g of tobacco hairy roots (Chahardoli et al, 2018). In addition, all of these plant-based production systems are based on stable transformation, which makes the process more complex and time consuming.…”

“…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.…”

“…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 (Bund o et al, 2014;Coca et al, 2006;Montesinos et al, 2016). To target AfpB to the apoplast or ER, we added the signal peptide of the tobacco AP24 protein (XP_009782398.1) to the amino terminus of the mature AfpB protein (Figure 2a).…”

Producción y utilización Biotecnológica de nuevas proteínas antifúngicas de hongos filamentosos

“…Despite the physicochemical properties of some of these peptides, several plant-based platforms have been reported to be suitable for the large-scale production of AMPs. Among them, there are chloroplasts-based or rice seeds-based production systems (Lee et al, 2001;Montesinos et al, 2016;Montesinos et al, 2017). The success of these strategies is based on limiting the accumulation of AMPs to storage organs in order to avoid interference with plant vegetative growth; or to limit it to specialized subcellular organelles that would both protect AMPs from protease degradation and facilitate their purification (Montesinos et al, 2016;Montesinos et al, 2017).…”