Multi-Omics Analysis Provides Insights into a Mosaic-Leaf Phenotype of Astaxanthin-Producing Tobacco

In metabolically engineered plants, the target products are usually uniformly distributed in the whole plant or specific tissues. When engineering tobacco to produce astaxanthin, a ketocarotenoid with strong antioxidant activity and multiple bioactivities, a scattered distribution of astaxanthin-pro...

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Main Authors: Jialin Wang, Zaifeng Du, Xiaoyang Lin, Peng Li, Shihao Sun, Changqing Yang, Yong Chen, Zhongfeng Zhang, Xue Yin, Ning Fang
Format: Article
Language:English
Published: MDPI AG 2025-03-01
Series:Plants
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Online Access:https://www.mdpi.com/2223-7747/14/6/965
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author Jialin Wang
Zaifeng Du
Xiaoyang Lin
Peng Li
Shihao Sun
Changqing Yang
Yong Chen
Zhongfeng Zhang
Xue Yin
Ning Fang
author_facet Jialin Wang
Zaifeng Du
Xiaoyang Lin
Peng Li
Shihao Sun
Changqing Yang
Yong Chen
Zhongfeng Zhang
Xue Yin
Ning Fang
author_sort Jialin Wang
collection DOAJ
description In metabolically engineered plants, the target products are usually uniformly distributed in the whole plant or specific tissues. When engineering tobacco to produce astaxanthin, a ketocarotenoid with strong antioxidant activity and multiple bioactivities, a scattered distribution of astaxanthin-producing regions was observed in a small portion of astaxanthin-producing tobacco plants, which caused mosaic-like red and green spots on the leaves (ASTA-mosaic). A physiological assay showed that the non-astaxanthin green region (Mosaic_G) had relatively higher chlorophyll content and better chloroplast structure than the astaxanthin-producing red region (Mosaic_R). Then, metabolomics, proteomics, and small RNA transcriptomics were employed to analyze the uneven distribution of astaxanthin-producing regions in tobacco leaves. The results of metabolomics and proteomics revealed a decrease in carotenoid metabolism, chlorophyll biosynthesis, and chlorophyll degradation in the Mosaic_G region. Pheophorbide a, an intermediate of chlorophyll degradation, was found to be significantly reduced in the Mosaic_G region, which was accompanied by the attenuation of chlorophyllase and pheophytinase, which catalyze the formation of pheophorbide a in chlorophyll degradation. Reductions in photosynthetic antenna proteins and photosystem-associated proteins were observed in the Mosaic_R region, consistent with the better chloroplast structure of the Mosaic_G region. Small RNA transcriptomics showed that several small RNAs could target chlorophyll-degradative genes, but they were more effective in targeting the astaxanthin biosynthetic genes. This finding was supported by the fact that the Mosaic_G region can remain green up to the senescence of tobacco leaves. This work provides insights into the mechanism of the uneven distribution of astaxanthin-producing regions in tobacco leaves and may contribute to the specialized utilization of tobacco plants for metabolic engineering.
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spelling doaj-art-787e70f2b80448aa9709a7c45b74ae3c2025-08-20T02:42:25ZengMDPI AGPlants2223-77472025-03-0114696510.3390/plants14060965Multi-Omics Analysis Provides Insights into a Mosaic-Leaf Phenotype of Astaxanthin-Producing TobaccoJialin Wang0Zaifeng Du1Xiaoyang Lin2Peng Li3Shihao Sun4Changqing Yang5Yong Chen6Zhongfeng Zhang7Xue Yin8Ning Fang9Key Laboratory of Synthetic Biology of Ministry of Agriculture and Rural Affairs, Tobacco Research Institute, Chinese Academy of Agricultural Sciences, Qingdao 266101, ChinaKey Laboratory of Synthetic Biology of Ministry of Agriculture and Rural Affairs, Tobacco Research Institute, Chinese Academy of Agricultural Sciences, Qingdao 266101, ChinaKey Laboratory of Synthetic Biology of Ministry of Agriculture and Rural Affairs, Tobacco Research Institute, Chinese Academy of Agricultural Sciences, Qingdao 266101, ChinaNo. 2 Department of Applied Research, Beijing Life Science Academy (BLSA), Beijing 102209, ChinaNo. 2 Department of Applied Research, Beijing Life Science Academy (BLSA), Beijing 102209, ChinaKey Laboratory of Synthetic Biology of Ministry of Agriculture and Rural Affairs, Tobacco Research Institute, Chinese Academy of Agricultural Sciences, Qingdao 266101, ChinaNo. 2 Department of Applied Research, Beijing Life Science Academy (BLSA), Beijing 102209, ChinaKey Laboratory of Synthetic Biology of Ministry of Agriculture and Rural Affairs, Tobacco Research Institute, Chinese Academy of Agricultural Sciences, Qingdao 266101, ChinaKey Laboratory of Synthetic Biology of Ministry of Agriculture and Rural Affairs, Tobacco Research Institute, Chinese Academy of Agricultural Sciences, Qingdao 266101, ChinaKey Laboratory of Synthetic Biology of Ministry of Agriculture and Rural Affairs, Tobacco Research Institute, Chinese Academy of Agricultural Sciences, Qingdao 266101, ChinaIn metabolically engineered plants, the target products are usually uniformly distributed in the whole plant or specific tissues. When engineering tobacco to produce astaxanthin, a ketocarotenoid with strong antioxidant activity and multiple bioactivities, a scattered distribution of astaxanthin-producing regions was observed in a small portion of astaxanthin-producing tobacco plants, which caused mosaic-like red and green spots on the leaves (ASTA-mosaic). A physiological assay showed that the non-astaxanthin green region (Mosaic_G) had relatively higher chlorophyll content and better chloroplast structure than the astaxanthin-producing red region (Mosaic_R). Then, metabolomics, proteomics, and small RNA transcriptomics were employed to analyze the uneven distribution of astaxanthin-producing regions in tobacco leaves. The results of metabolomics and proteomics revealed a decrease in carotenoid metabolism, chlorophyll biosynthesis, and chlorophyll degradation in the Mosaic_G region. Pheophorbide a, an intermediate of chlorophyll degradation, was found to be significantly reduced in the Mosaic_G region, which was accompanied by the attenuation of chlorophyllase and pheophytinase, which catalyze the formation of pheophorbide a in chlorophyll degradation. Reductions in photosynthetic antenna proteins and photosystem-associated proteins were observed in the Mosaic_R region, consistent with the better chloroplast structure of the Mosaic_G region. Small RNA transcriptomics showed that several small RNAs could target chlorophyll-degradative genes, but they were more effective in targeting the astaxanthin biosynthetic genes. This finding was supported by the fact that the Mosaic_G region can remain green up to the senescence of tobacco leaves. This work provides insights into the mechanism of the uneven distribution of astaxanthin-producing regions in tobacco leaves and may contribute to the specialized utilization of tobacco plants for metabolic engineering.https://www.mdpi.com/2223-7747/14/6/965astaxanthinmosaic-like spotschlorophyll metabolismmetabolomicsproteomicssmall RNA transcriptomics
spellingShingle Jialin Wang
Zaifeng Du
Xiaoyang Lin
Peng Li
Shihao Sun
Changqing Yang
Yong Chen
Zhongfeng Zhang
Xue Yin
Ning Fang
Multi-Omics Analysis Provides Insights into a Mosaic-Leaf Phenotype of Astaxanthin-Producing Tobacco
Plants
astaxanthin
mosaic-like spots
chlorophyll metabolism
metabolomics
proteomics
small RNA transcriptomics
title Multi-Omics Analysis Provides Insights into a Mosaic-Leaf Phenotype of Astaxanthin-Producing Tobacco
title_full Multi-Omics Analysis Provides Insights into a Mosaic-Leaf Phenotype of Astaxanthin-Producing Tobacco
title_fullStr Multi-Omics Analysis Provides Insights into a Mosaic-Leaf Phenotype of Astaxanthin-Producing Tobacco
title_full_unstemmed Multi-Omics Analysis Provides Insights into a Mosaic-Leaf Phenotype of Astaxanthin-Producing Tobacco
title_short Multi-Omics Analysis Provides Insights into a Mosaic-Leaf Phenotype of Astaxanthin-Producing Tobacco
title_sort multi omics analysis provides insights into a mosaic leaf phenotype of astaxanthin producing tobacco
topic astaxanthin
mosaic-like spots
chlorophyll metabolism
metabolomics
proteomics
small RNA transcriptomics
url https://www.mdpi.com/2223-7747/14/6/965
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