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1. Maize resistance to insect herbivory is enhanced by silencing expression of genes for jasmonate-isoleucine degradation using sugarcane mosaic virus

   Chung S, S. Z (June 2022, Plant direct)

   Previously, sugarcane mosaic virus (SCMV) was developed as a vector for transient expression of heterologous genes in Zea mays (maize). Here, we show that SCMV can also be applied for virus-induced gene silencing (VIGS) of endogenous maize genes. Comparison of sense and antisense VIGS constructs targeting maize phytoene desaturase (PDS) showed that antisense constructs resulted in a greater reduction in gene expression. In a time course of gene expression after infection with VIGS constructs targeting PDS, lesion mimic 22 (Les22), and Iodent japonica 1 (Ij1), efficient expression silencing was observed 2, 3, and 4 weeks after infection with SCMV. However, at Week 5, expression of Les22 and Ij1 was no longer significantly reduced compared with control plants. The defense signaling molecule jasmonate-isoleucine (JA-Ile) can be inactivated by 12C-hydroxylation and hydrolysis, and knockout of these genes leads to herbivore resistance. JA-Ile hydroxylases and hydrolases have been investigated in Arabidopsis, rice, and Nicotiana attenuata. To determine whether the maize homologs of these genes function in plant defense, we silenced expression of ZmCYP94B1 (predicted JA-Ile hydroxylase) and ZmJIH1 (predicted JA-Ile hydrolase) by VIGS with SCMV, which resulted in elevated expression of two defense-related genes, Maize Proteinase Inhibitor (MPI) and Ribosome Inactivating Protein 2 (RIP2). Although ZmCYP94B1 and ZmJIH1 gene expression silencing increased resistance to Spodoptera frugiperda (fall armyworm), Schistocerca americana (American birdwing grasshopper), and Rhopalosiphum maidis (corn leaf aphid), there was no additive effect from silencing the expression of both genes. Further work will be required to determine the more precise functions of these enzymes in regulating maize defenses. 

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2. Maize resistance to insect herbivory is enhanced by silencing expression of genes for jasmonate-isoleucine degradation using sugarcane mosaic virus

   Chung S, S. Z (June 2022, Plant direct)

   Previously, sugarcane mosaic virus (SCMV) was developed as a vector for transient expression of heterologous genes in Zea mays (maize). Here, we show that SCMV can also be applied for virus-induced gene silencing (VIGS) of endogenous maize genes. Comparison of sense and antisense VIGS constructs targeting maize phytoene desaturase (PDS) showed that antisense constructs resulted in a greater reduction in gene expression. In a time course of gene expression after infection with VIGS constructs targeting PDS, lesion mimic 22 (Les22), and Iodent japonica 1 (Ij1), efficient expression silencing was observed 2, 3, and 4 weeks after infection with SCMV. However, at Week 5, expression of Les22 and Ij1 was no longer significantly reduced compared with control plants. The defense signaling molecule jasmonate-isoleucine (JA-Ile) can be inactivated by 12C-hydroxylation and hydrolysis, and knockout of these genes leads to herbivore resistance. JA-Ile hydroxylases and hydrolases have been investigated in Arabidopsis, rice, and Nicotiana attenuata. To determine whether the maize homologs of these genes function in plant defense, we silenced expression of ZmCYP94B1 (predicted JA-Ile hydroxylase) and ZmJIH1 (predicted JA-Ile hydrolase) by VIGS with SCMV, which resulted in elevated expression of two defense-related genes, Maize Proteinase Inhibitor (MPI) and Ribosome Inactivating Protein 2 (RIP2). Although ZmCYP94B1 and ZmJIH1 gene expression silencing increased resistance to Spodoptera frugiperda (fall armyworm), Schistocerca americana (American birdwing grasshopper), and Rhopalosiphum maidis (corn leaf aphid), there was no additive effect from silencing the expression of both genes. Further work will be required to determine the more precise functions of these enzymes in regulating maize defenses. 

   more » « less
3. Maize resistance to insect herbivory is enhanced by silencing expression of genes for jasmonate-isoleucine degradation using sugarcane mosaic virus

   Chung S, S. Z (May 2022, Plant direct)

   Previously, sugarcane mosaic virus (SCMV) was developed as a vector for transient expression of heterologous genes in Zea mays (maize). Here, we show that SCMV can also be applied for virus-induced gene silencing (VIGS) of endogenous maize genes. Comparison of sense and antisense VIGS constructs targeting maize phytoene desaturase (PDS) showed that antisense constructs resulted in a greater reduction in gene expression. In a time course of gene expression after infection with VIGS con- structs targeting PDS, lesion mimic 22 (Les22), and Iodent japonica 1 (Ij1), efficient expression silencing was observed 2, 3, and 4 weeks after infection with SCMV. However, at Week 5, expression of Les22 and Ij1 was no longer significantly reduced compared with control plants. The defense signaling molecule jasmonate-isoleucine (JA-Ile) can be inactivated by 12C-hydroxylation and hydrolysis, and knockout of these genes leads to herbivore resistance. JA-Ile hydroxylases and hydrolases have been investigated in Arabidopsis, rice, and Nicotiana attenuata. To determine whether the maize homologs of these genes function in plant defense, we silenced expression of ZmCYP94B1 (predicted JA-Ile hydroxylase) and ZmJIH1 (predicted JA- Ile hydrolase) by VIGS with SCMV, which resulted in elevated expression of two defense-related genes, Maize Proteinase Inhibitor (MPI) and Ribosome Inactivating Pro- tein 2 (RIP2). Although ZmCYP94B1 and ZmJIH1 gene expression silencing increased resistance to Spodoptera frugiperda (fall armyworm), Schistocerca americana (American birdwing grasshopper), and Rhopalosiphum maidis (corn leaf aphid), there was no addi- tive effect from silencing the expression of both genes. Further work will be required to determine the more precise functions of these enzymes in regulating maize defenses. 

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4. Functional dissection of the ARGONAUTE7 promoter

   https://doi.org/10.1002/pld3.102  

   Hoyer, J. Steen; Pruneda‐Paz, Jose L.; Breton, Ghislain; Hassert, Mariah A.; Holcomb, Emily E.; Fowler, Halley; Bauer, Kaylyn M.; Mreen, Jacob; Kay, Steve A.; Carrington, James C. (January 2019, Plant Direct)

   Abstract ARGONAUTES are the central effector proteins ofRNAsilencing which bind target transcripts in a smallRNA‐guided manner.Arabidopsis thalianahas 10ARGONAUTE(AGO) genes, with specialized roles inRNA‐directedDNAmethylation, post‐transcriptional gene silencing, and antiviral defense. To better understand specialization amongAGOgenes at the level of transcriptional regulation we tested a library of 1497 transcription factors for binding to the promoters ofAGO1,AGO10, andAGO7using yeast 1‐hybrid assays. A ranked list of candidateDNA‐bindingTFs revealed binding of theAGO7promoter by a number of proteins in two families: the miR156‐regulatedSPLfamily and the miR319‐regulatedTCPfamily, both of which have roles in developmental timing and leaf morphology. Possible functions forSPLandTCPbinding are unclear: we showed that these binding sites are not required for the polar expression pattern ofAGO7, nor for the function ofAGO7in leaf shape. NormalAGO7transcription levels and function appear to depend instead on an adjacent 124‐bp region. Progress in understanding the structure of this promoter may aid efforts to understand how the conservedAGO7‐triggeredTAS3pathway functions in timing and polarity. 

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5. RNAi Strategies Against Downy Mildews: Insights Into dsRNA Uptake and Silencing

   https://doi.org/10.1111/mpp.70140  

   Göl, Deniz; Okechukwu, Emeka; Ünal, Gizem; Webb, Anne; Wood, Tom; Hong, Yiguo; Sherif, Sherif_M; Wacker, Theresa; Studholme, David_J; McDowell, John_M; et al (August 2025, Molecular Plant Pathology)

   ABSTRACT Downy mildew (DM) diseases are caused by destructive obligate pathogens with limited control options, posing a significant threat to global agriculture. RNA interference (RNAi) has emerged as a promising, environmentally sustainable strategy for disease management. We evaluated the efficacy of dsRNA‐mediated RNAi in suppressing key biological functions in DM pathogens ofArabidopsis thaliana, pea and lettuce:Hyaloperonospora arabidopsidis(Hpa),Peronospora viciaef. sp.pisi(Pvp) andBremia lactucae(Bl), respectively. Conserved genes,cellulose synthase 3(CesA3) andbeta‐tubulin(BTUB), were targeted. Silencing these genes significantly impaired spore germination and infection across species and reduced gene expression correlated with suppressed sporulation, confirming silencing efficacy. We tested dsRNAs from chemical synthesis, in vitro transcription, andEscherichia coliexpression. Uptake and silencing efficiency varied with dsRNA length and concentration. InHpa, short dsRNAs (21–25 bp) produced a variable spore germination rate, with 25 bp dsRNA causing a 247.10% increase, whereas longer dsRNAs (≥ 30 bp) completely inhibited germination. Similarly, inPvp, dsRNAs of 21–25 bp resulted in a 73.05%–77.46% germination rate, while 30–75 bp dsRNAs abolished germination. Confocal microscopy using Cy‐5‐labelled short‐synthesised dsRNA (SS‐dsRNA) confirmed uptake by spores. Sequence specificity influenced efficacy, highlighting the need for precise target design. Multiplexed RNAi impacted silencing synergistically, further reducing germination and sporulation inHpa. Importantly, SS‐dsRNA‐mediated silencing was durable, with reduced gene expression sustained at 4, 7, 10 and 11 days post‐inoculation. Taken together, our findings demonstrate the potential of dsRNA‐mediated gene silencing as a precise, sustainable tool for managing DM pathogens in multiple crop species. 

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