CRISPR-Cas9 mediated dsRNase knockout improves RNAi efficiency in the fall armyworm

Jinmo Koo; Guan Heng Zhu; Subba Reddy Palli

doi:10.1016/j.pestbp.2024.105839

CRISPR-Cas9 mediated dsRNase knockout improves RNAi efficiency in the fall armyworm

Jinmo Koo
, Guan Heng Zhu
, Subba Reddy Palli

Research output: Contribution to journal › Article › peer-review

16 Scopus citations

Abstract

Lepidopteran insects are refractory to RNA interference (RNAi) response, especially to orally delivered double-stranded RNA (dsRNA). High nuclease activity in the midgut lumen is proposed as one of the major reasons for RNAi insensitivity. We identified three dsRNase genes highly expressed in the midgut of fall armyworm (FAW), Spodoptera frugiperda. The genomic region harboring those three dsRNase genes was deleted using the CRISPR-Cas9-mediated genome editing method. A homozygous line with deletion of three dsRNase genes was produced. dsRNA degradation by midgut lumen contents of mutant larvae was lower than in wild-type larvae. Feeding dsRNA targeting the inhibitor of apoptosis (IAP) gene increased knockdown of the target gene and mortality in mutants compared to wild-type larvae. These results suggest that dsRNases in the midgut contribute to RNAi inefficiency in FAW. Formulations that protect dsRNA from dsRNase degradation may improve RNAi efficiency in FAW and other lepidopteran insects.

Original language	English
Article number	105839
Journal	Pesticide Biochemistry and Physiology
Volume	200
DOIs	https://doi.org/10.1016/j.pestbp.2024.105839
State	Published - Mar 2024

Bibliographical note

Publisher Copyright:
© 2023

Funding

The research reported in this publication was supported by Agriculture and Food Research Initiative Competitive Grant No. 2019-67013-29351 and the National Institute of Food and Agriculture and the US Department of Agriculture (under HATCH Project 2351177000 ). The content is solely the responsibility of the authors and does not necessarily represent the official views of the USDA.

Funders	Funder number
U.S. Department of Agriculture	2351177000
U.S. Department of Agriculture
US Department of Agriculture National Institute of Food and Agriculture, Agriculture and Food Research Initiative

Keywords

Genome editing
Midgut
Nuclease
Spodoptera frugiperda
dsRNA stability

ASJC Scopus subject areas

Agronomy and Crop Science
Health, Toxicology and Mutagenesis

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10.1016/j.pestbp.2024.105839

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title = "CRISPR-Cas9 mediated dsRNase knockout improves RNAi efficiency in the fall armyworm",

abstract = "Lepidopteran insects are refractory to RNA interference (RNAi) response, especially to orally delivered double-stranded RNA (dsRNA). High nuclease activity in the midgut lumen is proposed as one of the major reasons for RNAi insensitivity. We identified three dsRNase genes highly expressed in the midgut of fall armyworm (FAW), Spodoptera frugiperda. The genomic region harboring those three dsRNase genes was deleted using the CRISPR-Cas9-mediated genome editing method. A homozygous line with deletion of three dsRNase genes was produced. dsRNA degradation by midgut lumen contents of mutant larvae was lower than in wild-type larvae. Feeding dsRNA targeting the inhibitor of apoptosis (IAP) gene increased knockdown of the target gene and mortality in mutants compared to wild-type larvae. These results suggest that dsRNases in the midgut contribute to RNAi inefficiency in FAW. Formulations that protect dsRNA from dsRNase degradation may improve RNAi efficiency in FAW and other lepidopteran insects.",

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AU - Koo, Jinmo

AU - Zhu, Guan Heng

AU - Palli, Subba Reddy

PY - 2024/3

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N2 - Lepidopteran insects are refractory to RNA interference (RNAi) response, especially to orally delivered double-stranded RNA (dsRNA). High nuclease activity in the midgut lumen is proposed as one of the major reasons for RNAi insensitivity. We identified three dsRNase genes highly expressed in the midgut of fall armyworm (FAW), Spodoptera frugiperda. The genomic region harboring those three dsRNase genes was deleted using the CRISPR-Cas9-mediated genome editing method. A homozygous line with deletion of three dsRNase genes was produced. dsRNA degradation by midgut lumen contents of mutant larvae was lower than in wild-type larvae. Feeding dsRNA targeting the inhibitor of apoptosis (IAP) gene increased knockdown of the target gene and mortality in mutants compared to wild-type larvae. These results suggest that dsRNases in the midgut contribute to RNAi inefficiency in FAW. Formulations that protect dsRNA from dsRNase degradation may improve RNAi efficiency in FAW and other lepidopteran insects.

AB - Lepidopteran insects are refractory to RNA interference (RNAi) response, especially to orally delivered double-stranded RNA (dsRNA). High nuclease activity in the midgut lumen is proposed as one of the major reasons for RNAi insensitivity. We identified three dsRNase genes highly expressed in the midgut of fall armyworm (FAW), Spodoptera frugiperda. The genomic region harboring those three dsRNase genes was deleted using the CRISPR-Cas9-mediated genome editing method. A homozygous line with deletion of three dsRNase genes was produced. dsRNA degradation by midgut lumen contents of mutant larvae was lower than in wild-type larvae. Feeding dsRNA targeting the inhibitor of apoptosis (IAP) gene increased knockdown of the target gene and mortality in mutants compared to wild-type larvae. These results suggest that dsRNases in the midgut contribute to RNAi inefficiency in FAW. Formulations that protect dsRNA from dsRNase degradation may improve RNAi efficiency in FAW and other lepidopteran insects.

KW - Genome editing

KW - Midgut

KW - Nuclease

KW - Spodoptera frugiperda

KW - dsRNA stability

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SN - 0048-3575

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CRISPR-Cas9 mediated dsRNase knockout improves RNAi efficiency in the fall armyworm

Abstract

Bibliographical note

Funding

Keywords

ASJC Scopus subject areas

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