Bacterium-Mediated RNA Interference: Potential Application in Plant Protection

Goodfellow, Simon; Zhang, Daai; Wang, Ming-Bo; Zhang, Ren

doi:10.3390/plants8120572

Cited by 21 publications

(23 citation statements)

References 108 publications

(152 reference statements)

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“…Proper selection of the optimal physiological and environmental conditions is important to elicit efficient gene silencing in plants after foliar dsRNA application. According to Goodfellow et al [ 41 ], plant-associated endophytic bacteria have significant potential in respect of further development of exo-RNAi-based plant modifications. Thus, the development of the tools of exogenously induced RNAi could contribute to further achievements in crop improvement and to the implementation of new instruments in plant gene functional studies.…”

Section: Discussionmentioning

confidence: 99%

Physiological Conditions and dsRNA Application Approaches for Exogenously induced RNA Interference in Arabidopsis thaliana

Kiselev

Suprun

Aleynova

et al. 2021

Plants

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Recent studies have revealed that foliar application of double-stranded RNAs (dsRNAs) or small-interfering RNAs (siRNAs) encoding specific genes of plant pathogens triggered RNA interference (RNAi)-mediated silencing of the gene targets. However, a limited number of reports documented silencing of plant endogenes or transgenes after direct foliar RNA application. This study analyzed the importance of physiological conditions (plant age, time of day, soil moisture, high salinity, heat, and cold stresses) and different dsRNA application means (brush spreading, spraying, infiltration, inoculation, needle injection, and pipetting) for suppression of neomycin phosphotransferase II (NPTII) transgene in Arabidopsis thaliana, as transgenes are more prone to silencing. We observed a higher NPTII suppression when dsRNA was applied at late day period, being most efficient at night, which revealed a diurnal variation in dsRNA treatment efficacy. Exogenous NPTII-dsRNA considerably reduced NPTII expression in 4-week-old plants and only limited it in 2- and 6-week-old plants. In addition, a more discernible NPTII downregulation was detected under low soil moisture conditions. Treatment of adaxial and abaxial leaf surfaces by brushes, spraying, and pipetting showed a higher NPTII suppression, while infiltration and inoculation were less efficient. Thus, appropriate plant age, late time of day, low soil moisture, and optimal dsRNA application modes are important for exogenously induced gene silencing.

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Section: Discussionmentioning

confidence: 99%

Physiological Conditions and dsRNA Application Approaches for Exogenously induced RNA Interference in Arabidopsis thaliana

Kiselev

Suprun

Aleynova

et al. 2021

Plants

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“…Bacterium-mediated RNA interference, wherein live endophytic bacteria expressing dsRNA are applied to and colonize an organism to produce and facilitate the uptake of dsRNA, has been shown to be effective in insects [ 40 ]. This approach has been proposed as a potential improvement of RNAi in plants to overcome some of the limitations of more-traditional application strategies (spraying, infiltration and drenching), such as the duration and the systemic nature of the RNAi silencing effect [ 41 ].…”

Section: Discussionmentioning

confidence: 99%

Topical Application of Escherichia coli-Encapsulated dsRNA Induces Resistance in Nicotiana benthamiana to Potato Viruses and Involves RDR6 and Combined Activities of DCL2 and DCL4

Necira

Makki

Sanz-García

et al. 2021

Plants

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Exogenous application of double-stranded RNAs (dsRNAs) for inducing virus resistance in plants represents an attractive alternative to transgene-based silencing approaches. However, improvement of dsRNA stability in natural conditions is required in order to provide long-term protection against the targeted virus. Here, we tested the protective effect of topical application of Escherichia coli-encapsulated dsRNA compared to naked dsRNA against single and dual infection by Potato virus X expressing the green fluorescent protein (PVX-GFP) and Potato virus Y (PVY) in Nicotiana benthamiana. We found that, in our conditions, the effectiveness of E. coli-encapsulated dsRNA in providing RNAi-mediated protection did not differ from that of naked dsRNA. dsRNA vaccination was partly effective against a dual infection by PVX-GFP and PVY, manifested by a delay in the expression of the synergistic symptoms at early times after inoculation. Using PVX-GFP as a reporter virus together with a suite of RNAi knockdown transgenic lines, we have also shown that RNA-directed RNA polymerase 6 and the combined activities of DICER-like 2 (DCL2) and DCL4 act to promote efficient resistance to virus infection conferred by topical application of dsRNA in N. benthamiana. Our results provide evidence that exogenous dsRNA molecules are processed by the RNA silencing pathways commonly used by the host in response to virus infection.

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“…In this study, we exploited the systemic properties of dsRNAs and successfully utilized bacteria as an alternative to HIGS and SIGS methods to deliver RNAi to filamentous fungi. Bacteria have the advantage that they can be cultured in large volumes relatively inexpensively and protect the produced dsRNAs within their cells until their application [ 48 , 49 ]. Our results showed that even after bacterial lysis, the dsRNAs remained resistant to degradation for at least up to 24 h, and partially until 72 h, providing enough time for the dsRNAs to be absorbed by the fungal cells and induce silencing in target genes.…”

Section: Discussionmentioning

confidence: 99%

“…In our studies, we have exploited the widely used HT115(DE3) E. coli strain as the host bacterium for the production of dsRNAs [ 29 ], but other bacteria species could be potentially engineered to deliver RNAi to fungi, including for example bacteria used in biological control, plant endophytic or symbiotic bacteria, and several others [ 48 ]. Indeed, symbiotic gut bacteria of insects were recently successfully engineered to deliver RNAi to their insect hosts [ 50 ], indicating that the approach is amendable to different bacterial species that can be used to meet the needs of specific applications.…”

Section: Discussionmentioning

confidence: 99%

Targeted Delivery of Gene Silencing in Fungi Using Genetically Engineered Bacteria

Niño‐Sánchez

Chen

Souza

et al. 2021

JoF

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Exploiting RNA interference (RNAi) in disease control through non-transformative methods that overcome the hurdle of producing transgenic plants has attracted much attention over the last years. Here, we explored such a method and used non-pathogenic bacteria as a versatile system for delivering RNAi to fungi. Specifically, the RNaseIII-null mutant strain of Escherichia coli HT115(DE3) was transformed with two plasmid vectors that enabled the constitutive or IPTG-inducible production of double-stranded RNAs (dsRNAs) against genes involved in aflatoxins production in Aspergillus flavus (AflC) or virulence of Botrytis cinerea (BcSAS1). To facilitate the release of the dsRNAs, the bacterial cells were further genetically engineered to undergo a bacteriophage endolysin R-mediated autolysis, following a freeze-thaw cycle. Exposure under in vitro conditions of A. flavus or B. cinerea to living bacteria or their whole-cell autolysates induced silencing of AflC and BcSAS1 in a bacteria concentration-dependent manner, and instigated a reduction in aflatoxins production and mycelial growth, respectively. In planta applications of the living bacteria or their crude whole-cell autolysates produced similar results, thus creating a basis for translational research. These results demonstrate that bacteria can produce biologically active dsRNA against target genes in fungi and that bacteria-mediated RNAi can be used to control fungal pathogens.

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Bacterium-Mediated RNA Interference: Potential Application in Plant Protection

Cited by 21 publications

References 108 publications

Physiological Conditions and dsRNA Application Approaches for Exogenously induced RNA Interference in Arabidopsis thaliana

Physiological Conditions and dsRNA Application Approaches for Exogenously induced RNA Interference in Arabidopsis thaliana

Topical Application of Escherichia coli-Encapsulated dsRNA Induces Resistance in Nicotiana benthamiana to Potato Viruses and Involves RDR6 and Combined Activities of DCL2 and DCL4

Targeted Delivery of Gene Silencing in Fungi Using Genetically Engineered Bacteria

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