Silencing Dicer-Like Genes Reduces Virulence and sRNA Generation in <i>Penicillium italicum,</i> the Cause of Citrus Blue Mold
The Dicer protein is one of the most important components of RNAi machinery because it regulates the production of small RNAs (sRNAs) in eukaryotes. Here, Dicer1-like gene (Pit-DCL1) and Dicer2-like gene (Pit-DCL2) RNAi transformants were generated via pSilent-1 in <i>Penicillium italicum</...
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MDPI AG
2020-02-01
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| author | Chunxiao Yin Hong Zhu Yueming Jiang Yang Shan Liang Gong |
| author_facet | Chunxiao Yin Hong Zhu Yueming Jiang Yang Shan Liang Gong |
| author_sort | Chunxiao Yin |
| collection | DOAJ |
| description | The Dicer protein is one of the most important components of RNAi machinery because it regulates the production of small RNAs (sRNAs) in eukaryotes. Here, Dicer1-like gene (Pit-DCL1) and Dicer2-like gene (Pit-DCL2) RNAi transformants were generated via pSilent-1 in <i>Penicillium italicum</i> (Pit), which is the causal agent of citrus blue mold. Neither transformant showed a change in mycelial growth or sporulation ability, but the pathogenicity of the Pit-DCL2 RNAi transformant to citrus fruits was severely impaired, compared to that of the Pit-DCL1 RNAi transformant and the wild type. We further developed a citrus wound-mediated RNAi approach with a double-stranded fragment of Pit-DCL2 generated in vitro, which achieved an efficiency in reducing <i>Pi-Dcl2</i> expression and virulence that was similar to that of protoplast-mediated RNAi in <i>P. italicum,</i> suggesting that this approach is promising in the exogenous application of dsRNA to control pathogens on the surface of citrus fruits. In addition, sRNA sequencing revealed a total of 69.88 million potential sRNAs and 12 novel microRNA-like small RNAs (milRNAs), four of which have been predicated on target innate immunity or biotic stress-related genes in Valencia orange. These data suggest that both the Pit-DCL1 and Pit-DCL2 RNAi transformants severely disrupted the biogenesis of the potential milRNAs, which was further confirmed for some milRNAs by qRT-PCR or Northern blot analysis. These data suggest the sRNAs in <i>P. italicum</i> that may be involved in a molecular virulence mechanism termed cross-kingdom RNAi (ck-RNAi) by trafficking sRNA from <i>P. italicum</i> to citrus fruits. |
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| spelling | doaj.art-8ec35dfbdc8b42aea6321e7591e7ad592023-09-02T08:49:03ZengMDPI AGCells2073-44092020-02-019236310.3390/cells9020363cells9020363Silencing Dicer-Like Genes Reduces Virulence and sRNA Generation in <i>Penicillium italicum,</i> the Cause of Citrus Blue MoldChunxiao Yin0Hong Zhu1Yueming Jiang2Yang Shan3Liang Gong4Long Ping Branch, Graduate School of Hunan University, Changsha 410125, ChinaKey Laboratory of Plant Resource Conservation and Sustainable Utilization, Key Laboratory of Post-Harvest Handling of Fruits, Ministry of Agriculture, Guangdong Provincial Key Laboratory of Applied Botany, South China Botanical Garden, Chinese Academy of Sciences, Guangzhou 510650, ChinaKey Laboratory of Plant Resource Conservation and Sustainable Utilization, Key Laboratory of Post-Harvest Handling of Fruits, Ministry of Agriculture, Guangdong Provincial Key Laboratory of Applied Botany, South China Botanical Garden, Chinese Academy of Sciences, Guangzhou 510650, ChinaLong Ping Branch, Graduate School of Hunan University, Changsha 410125, ChinaKey Laboratory of Plant Resource Conservation and Sustainable Utilization, Key Laboratory of Post-Harvest Handling of Fruits, Ministry of Agriculture, Guangdong Provincial Key Laboratory of Applied Botany, South China Botanical Garden, Chinese Academy of Sciences, Guangzhou 510650, ChinaThe Dicer protein is one of the most important components of RNAi machinery because it regulates the production of small RNAs (sRNAs) in eukaryotes. Here, Dicer1-like gene (Pit-DCL1) and Dicer2-like gene (Pit-DCL2) RNAi transformants were generated via pSilent-1 in <i>Penicillium italicum</i> (Pit), which is the causal agent of citrus blue mold. Neither transformant showed a change in mycelial growth or sporulation ability, but the pathogenicity of the Pit-DCL2 RNAi transformant to citrus fruits was severely impaired, compared to that of the Pit-DCL1 RNAi transformant and the wild type. We further developed a citrus wound-mediated RNAi approach with a double-stranded fragment of Pit-DCL2 generated in vitro, which achieved an efficiency in reducing <i>Pi-Dcl2</i> expression and virulence that was similar to that of protoplast-mediated RNAi in <i>P. italicum,</i> suggesting that this approach is promising in the exogenous application of dsRNA to control pathogens on the surface of citrus fruits. In addition, sRNA sequencing revealed a total of 69.88 million potential sRNAs and 12 novel microRNA-like small RNAs (milRNAs), four of which have been predicated on target innate immunity or biotic stress-related genes in Valencia orange. These data suggest that both the Pit-DCL1 and Pit-DCL2 RNAi transformants severely disrupted the biogenesis of the potential milRNAs, which was further confirmed for some milRNAs by qRT-PCR or Northern blot analysis. These data suggest the sRNAs in <i>P. italicum</i> that may be involved in a molecular virulence mechanism termed cross-kingdom RNAi (ck-RNAi) by trafficking sRNA from <i>P. italicum</i> to citrus fruits.https://www.mdpi.com/2073-4409/9/2/363citrus<i>penicillium italicum</i>pathogenicityrna interferencecross-kingdom rnaidicersmall rna |
| spellingShingle | Chunxiao Yin Hong Zhu Yueming Jiang Yang Shan Liang Gong Silencing Dicer-Like Genes Reduces Virulence and sRNA Generation in <i>Penicillium italicum,</i> the Cause of Citrus Blue Mold Cells citrus <i>penicillium italicum</i> pathogenicity rna interference cross-kingdom rnai dicer small rna |
| title | Silencing Dicer-Like Genes Reduces Virulence and sRNA Generation in <i>Penicillium italicum,</i> the Cause of Citrus Blue Mold |
| title_full | Silencing Dicer-Like Genes Reduces Virulence and sRNA Generation in <i>Penicillium italicum,</i> the Cause of Citrus Blue Mold |
| title_fullStr | Silencing Dicer-Like Genes Reduces Virulence and sRNA Generation in <i>Penicillium italicum,</i> the Cause of Citrus Blue Mold |
| title_full_unstemmed | Silencing Dicer-Like Genes Reduces Virulence and sRNA Generation in <i>Penicillium italicum,</i> the Cause of Citrus Blue Mold |
| title_short | Silencing Dicer-Like Genes Reduces Virulence and sRNA Generation in <i>Penicillium italicum,</i> the Cause of Citrus Blue Mold |
| title_sort | silencing dicer like genes reduces virulence and srna generation in i penicillium italicum i the cause of citrus blue mold |
| topic | citrus <i>penicillium italicum</i> pathogenicity rna interference cross-kingdom rnai dicer small rna |
| url | https://www.mdpi.com/2073-4409/9/2/363 |
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