Off-target effects of RNAi correlate with the mismatch rate between dsRNA and non-target mRNA

Chen, Jiasheng; Peng, Yajun; Zhang, Hainan; Wang, Kangxu; Zhao, Chunqing; Zhu, Guan-Heng; Palli, Subba Reddy; Han, Zhaojun

doi:10.1080/15476286.2020.1868680

Cited by 66 publications

(101 citation statements)

References 65 publications

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“…This study adds to the conclusions from prior research on various insects that these effects tend to be mediated by high dsRNA concentrations [3,5,25], and it also demonstrates that dsRNA formulations can have significant antifeedant effects for orallydelivered RNAi. Second, there is a need to extend research on the relationship between the number of matching siRNA sequences and RNAi efficacy of dsRNA molecules [16,61,62], so that effective dsRNAs can be designed against single or closely-related pest species that vary in sequence identity with the target gene. Attention to these issues will facilitate the development and deployment of RNAi-based solutions for the management of plant sap-feeding and other insect pests.…”

Section: Discussionmentioning

confidence: 99%

“…However, these predictions of dsRNA specificity can be confounded by two issues. First, RNAi susceptibility to heterologous dsRNA can be obtained with a single matching 21-mer sequence, require several or many matching 21-mers, be achieved with as few as 15 contiguously matching bases and be tolerant of some sequence mismatches-varying with species, gene and dsRNA concentration [11,[15][16][17][18]. Second, animals, including insects, can respond to dsRNA in a sequence-non-specific manner, i.e., independent of the sequence of the dsRNA.…”

Section: Introductionmentioning

confidence: 99%

“…Second, animals, including insects, can respond to dsRNA in a sequence-non-specific manner, i.e., independent of the sequence of the dsRNA. In particular, various studies have shown that dsRNA with no matching 15-21-mer sequences in an insect genome can have substantial effects on antiviral immunity, gene expression and performance in insects [16,[19][20][21][22][23][24], although the molecular mechanisms are not fully understood. These effects are particularly pronounced for dsRNA administered at high concentrations, and it has been argued that high siRNA titers may saturate the core RNAi machinery [11,25].…”

Section: Introductionmentioning

confidence: 99%

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Non-Target Effects of dsRNA Molecules in Hemipteran Insects

Arora

Chung

Douglas

2021

Genes

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Insect pest control by RNA interference (RNAi)-mediated gene expression knockdown can be undermined by many factors, including small sequence differences between double-stranded RNA (dsRNA) and the target gene. It can also be compromised by effects that are independent of the dsRNA sequence on non-target organisms (known as sequence-non-specific effects). This study investigated the species-specificity of RNAi in plant sap-feeding hemipteran pests. We first demonstrated sequence-non-specific suppression of aphid feeding by dsRNA at dietary concentrations ≥0.5 µg µL−1. Then we quantified the expression of NUC (nuclease) genes in insects administered homologous dsRNA (with perfect sequence identity to the target species) or heterologous dsRNA (generated against a related gene of non-identical sequence in a different insect species). For the aphids Acyrthosiphon pisum and Myzus persicae, significantly reduced NUC expression was obtained with the homologous but not heterologous dsRNA at 0.2 µg µL−1, despite high dsNUC sequence identity. Follow-up experiments demonstrated significantly reduced expression of NUC genes in the whitefly Bemisia tabaci and mealybug Planococcus maritimus administered homologous dsNUCs, but not heterologous aphid dsNUCs. Our demonstration of inefficient expression knockdown by heterologous dsRNA in these insects suggests that maximal dsRNA sequence identity is required for RNAi targeting of related pest species, and that heterologous dsRNAs at appropriate concentrations may not be a major risk to non-target sap-feeding hemipterans.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Non-Target Effects of dsRNA Molecules in Hemipteran Insects

Arora

Chung

Douglas

2021

Genes

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“…With perfect sequence homology between dsRNA and mRNA only target gene suppression is expected, but it appears that siRNAs operate within cells with a certain level of "freedom" among targets. Mutation analyses showed that RNAi can be efficiently triggered with >80% sequence identity between siRNA and mRNA [158] but this mismatching tolerance can vary with insect species, target gene and dsRNA concentration [159,160]. Moreover, dsRNA can provoke responses independently of its sequence, affecting insect antiviral immunity, gene expression and performance [158,160].…”

Section: Rnai-related Concernsmentioning

confidence: 99%

“…Mutation analyses showed that RNAi can be efficiently triggered with >80% sequence identity between siRNA and mRNA [158] but this mismatching tolerance can vary with insect species, target gene and dsRNA concentration [159,160]. Moreover, dsRNA can provoke responses independently of its sequence, affecting insect antiviral immunity, gene expression and performance [158,160]. Although not fully understood, these effects are particularly pronounced for dsRNA administered at high concentrations, supposing that high levels of siRNA may saturate the core RNAi machinery [161].…”

Section: Rnai-related Concernsmentioning

confidence: 99%

Genetically Modified Potato for Pest Resistance: Thrift or Threat?

Raspor¹,

Cingel²

2021

Solanum Tuberosum - A Promising Crop for Starvation Problem

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Significant limitations in potato production are crop loss due to the damage made by insect pests, and the cost of enormous amount of chemicals, harmful to humans and environment, extensively used in their control. As an alternative, development of genetically modified potato offered possibility for pest management in a more sustainable, environmentally friendly way. Over the past 30 years introduction of pest resistance traits progressed from a single gene to multiple stacked events and from Bt-toxin expression to expression of proteins from non-Bt sources, dsRNA and their combination, while advances in molecular biology have brought “cleaner” gene manipulation technologies. However, together with benefits any new technology also bears its risks, and there are still a range of unanswered questions and concerns about long-term impact of genetically modified crops – that with knowledge and precautionary approaches can be avoided or mitigated. Sustainability of genetically modified crops for pest control largely depends on the willingness to gain and implement such knowledge.

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Inefficient uptake of small interfering RNAs is responsible for their inability to trigger RNA interference in Colorado potato beetle cells

Koo

Gurusamy

Palli

2023

Arch Insect Biochem Physiol

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There has been limited success in the usage of exogenous small interference RNA (siRNA) or small hairpin RNA (shRNA) to trigger RNA interference (RNAi) in insects. Instead, long double‐stranded RNAs (dsRNA) are used to induce knockdown of target genes in insects. Here, we compared the potency of si/sh RNAs and dsRNA in Colorado potato beetle (CPB) cells. CPB cells showed highly efficient RNAi response to dsRNA. However, si/sh RNAs were inefficient in triggering RNAi in CPB cells. Confocal microscopy observations of Cy3 labeled‐si/sh RNA cellular uptake revealed reduced si/sh RNA uptake compared to dsRNA. si/sh RNAs were stable in the conditioned media of CPB cells. Although in a small amount, when internalized by CPB cells, the si/sh RNAs were processed by the Dicer enzyme. Lipid‐mediated transfection and chimeric dsRNA approaches were used to improve the delivery of si/sh RNAs. Our results suggest that the uptake of si/sh RNAs is inefficient in CPB cells, resulting in ineffective RNAi response. However, with the help of effective delivery methods, si/sh RNA could be a useful option for developing target‐specific RNAi‐mediated biopesticides.

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Off-target effects of RNAi correlate with the mismatch rate between dsRNA and non-target mRNA

Cited by 66 publications

References 65 publications

Non-Target Effects of dsRNA Molecules in Hemipteran Insects

Non-Target Effects of dsRNA Molecules in Hemipteran Insects

Genetically Modified Potato for Pest Resistance: Thrift or Threat?

Inefficient uptake of small interfering RNAs is responsible for their inability to trigger RNA interference in Colorado potato beetle cells

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