I-<i>Sce</i>I Endonuclease-Mediated Plant Genome Editing by Protein Transport through a Bacterial Type III Secretion System
<i>Xanthomonas campestris</i> is one of bacteria carrying a type III secretion system which transports their effector proteins into host plant cells to disturb host defense system for their infection. To establish a genome editing system without introducing any foreign gene, we attempted...
| Main Authors: | , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2020-08-01
|
| Series: | Plants |
| Subjects: | |
| Online Access: | https://www.mdpi.com/2223-7747/9/9/1070 |
| _version_ | 1797556654071349248 |
|---|---|
| author | Yuki Yanagawa Kasumi Takeuchi Masaki Endo Ayako Furutani Hirokazu Ochiai Seiichi Toki Ichiro Mitsuhara |
| author_facet | Yuki Yanagawa Kasumi Takeuchi Masaki Endo Ayako Furutani Hirokazu Ochiai Seiichi Toki Ichiro Mitsuhara |
| author_sort | Yuki Yanagawa |
| collection | DOAJ |
| description | <i>Xanthomonas campestris</i> is one of bacteria carrying a type III secretion system which transports their effector proteins into host plant cells to disturb host defense system for their infection. To establish a genome editing system without introducing any foreign gene, we attempted to introduce genome editing enzymes through the type III secretion system. In a test of protein transfer, <i>X. campestris</i> pv. <i>campestris</i> (<i>Xcc</i>) transported a considerable amount of a reporter protein sGFP-CyaA into tobacco plant cells under the control of the type III secretion system while maintaining cell viability. For proof of concept for genome editing, we used a reporter tobacco plant containing a luciferase (LUC) gene interrupted by a meganuclease I-<i>Sce</i>I recognition sequence; this plant exhibits chemiluminescence of LUC only when a frameshift mutation is introduced at the I-<i>Sce</i>I recognition site. Luciferase signal was observed in tobacco leaves infected by <i>Xcc</i> carrying an <i>I-SceI</i> gene which secretes I-<i>Sce</i>I protein through the type III system, but not leaves infected by <i>Xcc</i> carrying a vector control. Genome-edited tobacco plant could be regenerated from a piece of infected leaf piece by repeated selection of LUC positive calli. Sequence analysis revealed that the regenerated tobacco plant possessed a base deletion in the I-<i>Sce</i>I recognition sequence that activated the <i>LUC</i> gene, indicating genome editing by I-<i>Sce</i>I protein transferred through the type III secretion system of <i>Xcc</i>. |
| first_indexed | 2024-03-10T17:05:55Z |
| format | Article |
| id | doaj.art-688f6a61b88b4030a8d9ac6d23304ab3 |
| institution | Directory Open Access Journal |
| issn | 2223-7747 |
| language | English |
| last_indexed | 2024-03-10T17:05:55Z |
| publishDate | 2020-08-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Plants |
| spelling | doaj.art-688f6a61b88b4030a8d9ac6d23304ab32023-11-20T10:48:56ZengMDPI AGPlants2223-77472020-08-0199107010.3390/plants9091070I-<i>Sce</i>I Endonuclease-Mediated Plant Genome Editing by Protein Transport through a Bacterial Type III Secretion SystemYuki Yanagawa0Kasumi Takeuchi1Masaki Endo2Ayako Furutani3Hirokazu Ochiai4Seiichi Toki5Ichiro Mitsuhara6Institute of Agrobiological Sciences, NARO, 1-2 Owashi, Tsukuba, Ibaraki 305-8634, JapanInstitute of Agrobiological Sciences, NARO, 1-2 Owashi, Tsukuba, Ibaraki 305-8634, JapanInstitute of Agrobiological Sciences, NARO, 1-2 Owashi, Tsukuba, Ibaraki 305-8634, JapanGene Research Center, Ibaraki University, 3-21-1 Ami, Inashiki, Ibaraki 300-0393, JapanInstitute of Agrobiological Sciences, NARO, 1-2 Owashi, Tsukuba, Ibaraki 305-8634, JapanInstitute of Agrobiological Sciences, NARO, 1-2 Owashi, Tsukuba, Ibaraki 305-8634, JapanInstitute of Agrobiological Sciences, NARO, 1-2 Owashi, Tsukuba, Ibaraki 305-8634, Japan<i>Xanthomonas campestris</i> is one of bacteria carrying a type III secretion system which transports their effector proteins into host plant cells to disturb host defense system for their infection. To establish a genome editing system without introducing any foreign gene, we attempted to introduce genome editing enzymes through the type III secretion system. In a test of protein transfer, <i>X. campestris</i> pv. <i>campestris</i> (<i>Xcc</i>) transported a considerable amount of a reporter protein sGFP-CyaA into tobacco plant cells under the control of the type III secretion system while maintaining cell viability. For proof of concept for genome editing, we used a reporter tobacco plant containing a luciferase (LUC) gene interrupted by a meganuclease I-<i>Sce</i>I recognition sequence; this plant exhibits chemiluminescence of LUC only when a frameshift mutation is introduced at the I-<i>Sce</i>I recognition site. Luciferase signal was observed in tobacco leaves infected by <i>Xcc</i> carrying an <i>I-SceI</i> gene which secretes I-<i>Sce</i>I protein through the type III system, but not leaves infected by <i>Xcc</i> carrying a vector control. Genome-edited tobacco plant could be regenerated from a piece of infected leaf piece by repeated selection of LUC positive calli. Sequence analysis revealed that the regenerated tobacco plant possessed a base deletion in the I-<i>Sce</i>I recognition sequence that activated the <i>LUC</i> gene, indicating genome editing by I-<i>Sce</i>I protein transferred through the type III secretion system of <i>Xcc</i>.https://www.mdpi.com/2223-7747/9/9/1070bacterial type III secretion system<i>Xanthomonas campestris</i>I-<i>Sce</i>Iprotein transportationgenome editing |
| spellingShingle | Yuki Yanagawa Kasumi Takeuchi Masaki Endo Ayako Furutani Hirokazu Ochiai Seiichi Toki Ichiro Mitsuhara I-<i>Sce</i>I Endonuclease-Mediated Plant Genome Editing by Protein Transport through a Bacterial Type III Secretion System Plants bacterial type III secretion system <i>Xanthomonas campestris</i> I-<i>Sce</i>I protein transportation genome editing |
| title | I-<i>Sce</i>I Endonuclease-Mediated Plant Genome Editing by Protein Transport through a Bacterial Type III Secretion System |
| title_full | I-<i>Sce</i>I Endonuclease-Mediated Plant Genome Editing by Protein Transport through a Bacterial Type III Secretion System |
| title_fullStr | I-<i>Sce</i>I Endonuclease-Mediated Plant Genome Editing by Protein Transport through a Bacterial Type III Secretion System |
| title_full_unstemmed | I-<i>Sce</i>I Endonuclease-Mediated Plant Genome Editing by Protein Transport through a Bacterial Type III Secretion System |
| title_short | I-<i>Sce</i>I Endonuclease-Mediated Plant Genome Editing by Protein Transport through a Bacterial Type III Secretion System |
| title_sort | i i sce i i endonuclease mediated plant genome editing by protein transport through a bacterial type iii secretion system |
| topic | bacterial type III secretion system <i>Xanthomonas campestris</i> I-<i>Sce</i>I protein transportation genome editing |
| url | https://www.mdpi.com/2223-7747/9/9/1070 |
| work_keys_str_mv | AT yukiyanagawa iisceiiendonucleasemediatedplantgenomeeditingbyproteintransportthroughabacterialtypeiiisecretionsystem AT kasumitakeuchi iisceiiendonucleasemediatedplantgenomeeditingbyproteintransportthroughabacterialtypeiiisecretionsystem AT masakiendo iisceiiendonucleasemediatedplantgenomeeditingbyproteintransportthroughabacterialtypeiiisecretionsystem AT ayakofurutani iisceiiendonucleasemediatedplantgenomeeditingbyproteintransportthroughabacterialtypeiiisecretionsystem AT hirokazuochiai iisceiiendonucleasemediatedplantgenomeeditingbyproteintransportthroughabacterialtypeiiisecretionsystem AT seiichitoki iisceiiendonucleasemediatedplantgenomeeditingbyproteintransportthroughabacterialtypeiiisecretionsystem AT ichiromitsuhara iisceiiendonucleasemediatedplantgenomeeditingbyproteintransportthroughabacterialtypeiiisecretionsystem |