Cargando…

Isolation of uracil auxotroph mutants of coral symbiont alga for symbiosis studies

Coral reef ecosystems rely on stable symbiotic relationship between the dinoflagellate Symbiodinium spp. and host cnidarian animals. The collapse of such symbiosis could cause coral ‘bleaching’ and subsequent host death. Despite huge interest on Symbiodinium, lack of mutant strains and readily avail...

Descripción completa

Detalles Bibliográficos
Autores principales: Ishii, Yuu, Maruyama, Shinichiro, Fujimura-Kamada, Konomi, Kutsuna, Natsumaro, Takahashi, Shunichi, Kawata, Masakado, Minagawa, Jun
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group UK 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5818653/
https://www.ncbi.nlm.nih.gov/pubmed/29459692
http://dx.doi.org/10.1038/s41598-018-21499-3
_version_ 1783301068715720704
author Ishii, Yuu
Maruyama, Shinichiro
Fujimura-Kamada, Konomi
Kutsuna, Natsumaro
Takahashi, Shunichi
Kawata, Masakado
Minagawa, Jun
author_facet Ishii, Yuu
Maruyama, Shinichiro
Fujimura-Kamada, Konomi
Kutsuna, Natsumaro
Takahashi, Shunichi
Kawata, Masakado
Minagawa, Jun
author_sort Ishii, Yuu
collection PubMed
description Coral reef ecosystems rely on stable symbiotic relationship between the dinoflagellate Symbiodinium spp. and host cnidarian animals. The collapse of such symbiosis could cause coral ‘bleaching’ and subsequent host death. Despite huge interest on Symbiodinium, lack of mutant strains and readily available genetic tools have hampered molecular research. A major issue was the tolerance to marker antibiotics. Here, we isolated Symbiodinium mutants requiring uracil for growth, and hence, useful in transformation screening. We cultured Symbiodinium spp. cells in the presence of 5-fluoroorotic acid (5FOA), which inhibits the growth of cells expressing URA3 encoding orotidine-5′-monophosphate decarboxylase, and isolated cells that require uracil for growth. Sequence analyses and genetic complementation tests using yeast demonstrated that one of the mutant cell lines had a point mutation in URA3, resulting in a splicing error at an unusual exon–intron junction, and consequently, loss of enzyme activity. This mutant could maintain a symbiotic relationship with the model sea anemone Exaiptasia pallida only in sea water containing uracil. Results show that the URA3 mutant will be a useful tool for screening Symbiodinium transformants, both ex and in hospite, as survival in the absence of uracil is possible only upon successful introduction of URA3.
format Online
Article
Text
id pubmed-5818653
institution National Center for Biotechnology Information
language English
publishDate 2018
publisher Nature Publishing Group UK
record_format MEDLINE/PubMed
spelling pubmed-58186532018-02-26 Isolation of uracil auxotroph mutants of coral symbiont alga for symbiosis studies Ishii, Yuu Maruyama, Shinichiro Fujimura-Kamada, Konomi Kutsuna, Natsumaro Takahashi, Shunichi Kawata, Masakado Minagawa, Jun Sci Rep Article Coral reef ecosystems rely on stable symbiotic relationship between the dinoflagellate Symbiodinium spp. and host cnidarian animals. The collapse of such symbiosis could cause coral ‘bleaching’ and subsequent host death. Despite huge interest on Symbiodinium, lack of mutant strains and readily available genetic tools have hampered molecular research. A major issue was the tolerance to marker antibiotics. Here, we isolated Symbiodinium mutants requiring uracil for growth, and hence, useful in transformation screening. We cultured Symbiodinium spp. cells in the presence of 5-fluoroorotic acid (5FOA), which inhibits the growth of cells expressing URA3 encoding orotidine-5′-monophosphate decarboxylase, and isolated cells that require uracil for growth. Sequence analyses and genetic complementation tests using yeast demonstrated that one of the mutant cell lines had a point mutation in URA3, resulting in a splicing error at an unusual exon–intron junction, and consequently, loss of enzyme activity. This mutant could maintain a symbiotic relationship with the model sea anemone Exaiptasia pallida only in sea water containing uracil. Results show that the URA3 mutant will be a useful tool for screening Symbiodinium transformants, both ex and in hospite, as survival in the absence of uracil is possible only upon successful introduction of URA3. Nature Publishing Group UK 2018-02-19 /pmc/articles/PMC5818653/ /pubmed/29459692 http://dx.doi.org/10.1038/s41598-018-21499-3 Text en © The Author(s) 2018 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
spellingShingle Article
Ishii, Yuu
Maruyama, Shinichiro
Fujimura-Kamada, Konomi
Kutsuna, Natsumaro
Takahashi, Shunichi
Kawata, Masakado
Minagawa, Jun
Isolation of uracil auxotroph mutants of coral symbiont alga for symbiosis studies
title Isolation of uracil auxotroph mutants of coral symbiont alga for symbiosis studies
title_full Isolation of uracil auxotroph mutants of coral symbiont alga for symbiosis studies
title_fullStr Isolation of uracil auxotroph mutants of coral symbiont alga for symbiosis studies
title_full_unstemmed Isolation of uracil auxotroph mutants of coral symbiont alga for symbiosis studies
title_short Isolation of uracil auxotroph mutants of coral symbiont alga for symbiosis studies
title_sort isolation of uracil auxotroph mutants of coral symbiont alga for symbiosis studies
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5818653/
https://www.ncbi.nlm.nih.gov/pubmed/29459692
http://dx.doi.org/10.1038/s41598-018-21499-3
work_keys_str_mv AT ishiiyuu isolationofuracilauxotrophmutantsofcoralsymbiontalgaforsymbiosisstudies
AT maruyamashinichiro isolationofuracilauxotrophmutantsofcoralsymbiontalgaforsymbiosisstudies
AT fujimurakamadakonomi isolationofuracilauxotrophmutantsofcoralsymbiontalgaforsymbiosisstudies
AT kutsunanatsumaro isolationofuracilauxotrophmutantsofcoralsymbiontalgaforsymbiosisstudies
AT takahashishunichi isolationofuracilauxotrophmutantsofcoralsymbiontalgaforsymbiosisstudies
AT kawatamasakado isolationofuracilauxotrophmutantsofcoralsymbiontalgaforsymbiosisstudies
AT minagawajun isolationofuracilauxotrophmutantsofcoralsymbiontalgaforsymbiosisstudies