Retrotransposon and CRISPR/Cas9‑mediated knockout of NOD26 impairs the legume‑rhizobia symbiosis
The multifunctional channel NOD26, identified and extensively studied (both biochemically and biophysically) in soybean, is a major protein component of the symbiosome membrane. The water and ammonia transport activities of NOD26 are thought to be important for nodule development, osmotic balance, a...
| Autores principales: | , , , , , , , , , , , |
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| Formato: | info:ar-repo/semantics/artículo |
| Lenguaje: | Inglés |
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Springer
2022
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| Materias: | |
| Acceso en línea: | http://hdl.handle.net/20.500.12123/12639 https://link.springer.com/article/10.1007/s11240-022-02357-7 https://doi.org/10.1007/s11240-022-02357-7 |
| _version_ | 1855036799644073984 |
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| author | Frare, Romina Alejandra Stritzler, Margarita Gomez, Maria Cristina Tajima, Hiromi Pascuan, Cecilia Gabriela López Fernández, María Paula Bottero, Ana Emilia Nikel, Pablo I. Alleva, Karina Edith Ayub, Nicolás Daniel Blumwald, Eduardo Soto, Gabriela Cynthia |
| author_browse | Alleva, Karina Edith Ayub, Nicolás Daniel Blumwald, Eduardo Bottero, Ana Emilia Frare, Romina Alejandra Gomez, Maria Cristina López Fernández, María Paula Nikel, Pablo I. Pascuan, Cecilia Gabriela Soto, Gabriela Cynthia Stritzler, Margarita Tajima, Hiromi |
| author_facet | Frare, Romina Alejandra Stritzler, Margarita Gomez, Maria Cristina Tajima, Hiromi Pascuan, Cecilia Gabriela López Fernández, María Paula Bottero, Ana Emilia Nikel, Pablo I. Alleva, Karina Edith Ayub, Nicolás Daniel Blumwald, Eduardo Soto, Gabriela Cynthia |
| author_sort | Frare, Romina Alejandra |
| collection | INTA Digital |
| description | The multifunctional channel NOD26, identified and extensively studied (both biochemically and biophysically) in soybean, is a major protein component of the symbiosome membrane. The water and ammonia transport activities of NOD26 are thought to be important for nodule development, osmotic balance, and ammonia efflux from the symbiosome. However, the widely accepted relevance of NOD26 in nitrogen-fixing symbiosis has never been explored in planta. Recently, we have reported the emergence of NOD26 in the nitrogen-fixing clade of angiosperms via tandem duplication. Here, we characterized the two copies of NOD26 from Medicago truncatula (Medtr8g087710 and Medtr8g087720) in their transport abilities, and at gene expression and genetic levels. Similar to their homologous soybean gene, MtNOD26 genes encode water and ammonia transport activities in heterologous expression systems. By using multiple transcriptional studies (RT-qPCR, transcriptional fusion and RNA-Seq analyses), we found that the expression of MtNOD26 copies is restricted to the nodule and gradually increases from the bacteria-free meristematic region to the nitrogen-fixation zone. Under nitrogen-limiting soil conditions, the homozygous insertional mutant lines of these two MtNOD26 genes had the same aberrant nodulation phenotype and chlorosis. Similar to uninoculated wild-type plants, inoculated mutants were unable to grow in minimal medium without a nitrogen source. Using the CRISPR/Cas9 system, we have edited the orthologous NOD26 genes in Medicago sativa (alfalfa), generating plants with aberrant nodules, chlorosis and impaired grow under nitrogen-limiting conditions. Collectively, our findings suggest functional equivalence between NOD26 copies and underline a crucial role of NOD26 in symbiotic nitrogen fixation. |
| format | info:ar-repo/semantics/artículo |
| id | INTA12639 |
| institution | Instituto Nacional de Tecnología Agropecuaria (INTA -Argentina) |
| language | Inglés |
| publishDate | 2022 |
| publishDateRange | 2022 |
| publishDateSort | 2022 |
| publisher | Springer |
| publisherStr | Springer |
| record_format | dspace |
| spelling | INTA126392022-08-22T10:20:12Z Retrotransposon and CRISPR/Cas9‑mediated knockout of NOD26 impairs the legume‑rhizobia symbiosis Frare, Romina Alejandra Stritzler, Margarita Gomez, Maria Cristina Tajima, Hiromi Pascuan, Cecilia Gabriela López Fernández, María Paula Bottero, Ana Emilia Nikel, Pablo I. Alleva, Karina Edith Ayub, Nicolás Daniel Blumwald, Eduardo Soto, Gabriela Cynthia Symbiosis Legumes Rhizobiaceae Ammonia CRISPR Simbiosis Leguminosas Amoníaco Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Interespaciadas The multifunctional channel NOD26, identified and extensively studied (both biochemically and biophysically) in soybean, is a major protein component of the symbiosome membrane. The water and ammonia transport activities of NOD26 are thought to be important for nodule development, osmotic balance, and ammonia efflux from the symbiosome. However, the widely accepted relevance of NOD26 in nitrogen-fixing symbiosis has never been explored in planta. Recently, we have reported the emergence of NOD26 in the nitrogen-fixing clade of angiosperms via tandem duplication. Here, we characterized the two copies of NOD26 from Medicago truncatula (Medtr8g087710 and Medtr8g087720) in their transport abilities, and at gene expression and genetic levels. Similar to their homologous soybean gene, MtNOD26 genes encode water and ammonia transport activities in heterologous expression systems. By using multiple transcriptional studies (RT-qPCR, transcriptional fusion and RNA-Seq analyses), we found that the expression of MtNOD26 copies is restricted to the nodule and gradually increases from the bacteria-free meristematic region to the nitrogen-fixation zone. Under nitrogen-limiting soil conditions, the homozygous insertional mutant lines of these two MtNOD26 genes had the same aberrant nodulation phenotype and chlorosis. Similar to uninoculated wild-type plants, inoculated mutants were unable to grow in minimal medium without a nitrogen source. Using the CRISPR/Cas9 system, we have edited the orthologous NOD26 genes in Medicago sativa (alfalfa), generating plants with aberrant nodules, chlorosis and impaired grow under nitrogen-limiting conditions. Collectively, our findings suggest functional equivalence between NOD26 copies and underline a crucial role of NOD26 in symbiotic nitrogen fixation. Instituto de Biotecnología Fil: Frare, Romina Alejandra. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular; Argentina Fil: Frare, Romina Alejandra. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Genética Ewald A. Favret; Argentina Fil: Frare, Romina Alejandra. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Stritzler, Margarita. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular; Argentina Fil: Stritzler, Margarita. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Genética Ewald A. Favret; Argentina Fil: Stritzler, Margarita. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Gomez, Maria Cristina. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular; Argentina Fil: Gomez, Maria Cristina. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Genética Ewald A. Favret; Argentina Fil: Gomez, Maria Cristina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Tajima, Hiromi. University of California. Department of Plant Sciences; Estados Unidos Fil: Pascuan, Cecilia Gabriela. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular; Argentina Fil: Pascuan, Cecilia Gabriela. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Genética Ewald A. Favret; Argentina Fil: Pascuan, Cecilia Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: López Fernández, María Paula. Universidad Nacional de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina Fil: Bottero, Ana Emilia. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular; Argentina Fil: Bottero, Ana Emilia. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Genética Ewald A. Favret; Argentina Fil: Bottero, Ana Emilia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Nikel, Pablo I. Technical University of Denmark. The Novo Nordisk Foundation Center for Biosustainability; Dinamarca Fil: Alleva, Karina Edith. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Fisicoquímica Biológica; Argentina Fil: Ayub, Nicolás Daniel. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular; Argentina Fil: Ayub, Nicolás Daniel. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Genética Ewald A. Favret; Argentina Fil: Ayub, Nicolás Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Blumwald, Eduardo. University of California. Department of Plant Sciences; Estados Unidos Fil: Soto, Gabriela Cynthia. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular; Argentina Fil: Soto, Gabriela Cynthia. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Genética Ewald A. Favret; Argentina Fil: Soto, Gabriela Cynthia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina 2022-08-22T10:16:18Z 2022-08-22T10:16:18Z 2022-07 info:ar-repo/semantics/artículo info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion http://hdl.handle.net/20.500.12123/12639 https://link.springer.com/article/10.1007/s11240-022-02357-7 1573-5044 https://doi.org/10.1007/s11240-022-02357-7 eng info:eu-repograntAgreement/INTA/2019-PE-E6-I115-001/2019-PE-E6-I115-001/AR./Edición génica, transgénesis y mutagénesis como generadores de nueva variabilidad en especies de interés agropecuario info:eu-repograntAgreement/INTA/2019-PD-E6-I116-001/2019-PD-E6-I116-001/AR./Identificación y análisis funcional de genes o redes génicas de interés biotecnológico con fin agropecuario, forestal, agroalimentario y/o agroindustrial. info:eu-repo/semantics/restrictedAccess application/pdf Springer Plant Cell, Tissue and Organ Culture (Published: 27 July 2022) |
| spellingShingle | Symbiosis Legumes Rhizobiaceae Ammonia CRISPR Simbiosis Leguminosas Amoníaco Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Interespaciadas Frare, Romina Alejandra Stritzler, Margarita Gomez, Maria Cristina Tajima, Hiromi Pascuan, Cecilia Gabriela López Fernández, María Paula Bottero, Ana Emilia Nikel, Pablo I. Alleva, Karina Edith Ayub, Nicolás Daniel Blumwald, Eduardo Soto, Gabriela Cynthia Retrotransposon and CRISPR/Cas9‑mediated knockout of NOD26 impairs the legume‑rhizobia symbiosis |
| title | Retrotransposon and CRISPR/Cas9‑mediated knockout of NOD26 impairs the legume‑rhizobia symbiosis |
| title_full | Retrotransposon and CRISPR/Cas9‑mediated knockout of NOD26 impairs the legume‑rhizobia symbiosis |
| title_fullStr | Retrotransposon and CRISPR/Cas9‑mediated knockout of NOD26 impairs the legume‑rhizobia symbiosis |
| title_full_unstemmed | Retrotransposon and CRISPR/Cas9‑mediated knockout of NOD26 impairs the legume‑rhizobia symbiosis |
| title_short | Retrotransposon and CRISPR/Cas9‑mediated knockout of NOD26 impairs the legume‑rhizobia symbiosis |
| title_sort | retrotransposon and crispr cas9 mediated knockout of nod26 impairs the legume rhizobia symbiosis |
| topic | Symbiosis Legumes Rhizobiaceae Ammonia CRISPR Simbiosis Leguminosas Amoníaco Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Interespaciadas |
| url | http://hdl.handle.net/20.500.12123/12639 https://link.springer.com/article/10.1007/s11240-022-02357-7 https://doi.org/10.1007/s11240-022-02357-7 |
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