Heterologous expression of rice 9-cis-epoxycarotenoid dioxygenase 4 (OsNCED4) in Arabidopsis confers sugar oversensitivity and drought tolerance

Botanical Studies, Jan 2018

Background The 9-cis-epoxycarotenoid dioxygenases OsNCED4 was cloned from rice in conjunction with OsNCED 1-3 and 5, of which 3 has been shown to function in ABA biosynthesis and alteration of leaf morphology. In higher plants, NCEDs have been shown to be key enzymes controlling ABA biosynthesis and belong to a differentially expressed gene family. Aside from OsNCED3, it remains largely unknown if other OsNCED genes are involved in ABA biosynthesis in rice. Thus, transgenic Arabidopsis plants overexpressing OsNCED4 were generated in the 129B08/nced3 mutant background to explore OsNCED4 function in ABA biosynthesis. Results Heterologous expression of OsNCED4 in Arabidopsis increased ABA levels and altered plant size and leaf shape, delayed seed germination, caused sugar oversensitivity in post-germination growth, and enhanced tolerance to drought. The native OsNCED3 and OsNCED4 promoters were expressed in an overlapping pattern in rice seeds and young seedlings, suggesting possible functional redundancy between OsNCED3 and OsNCED4. At the one-leaf stage, similar regulation of OsNCED3 and OsNCED4 gene expression in roots or leaves in response to moderate salt stress (150 mM NaCl) was observed. Conclusion Like OsNCED3, OsNCED4 is functionally active in ABA biosynthesis in rice. OsNCED3 and OsNCED4 might play redundant roles in controlling ABA biosynthesis in rice, as suggested by GUS staining assay, but this should be further analyzed through complementation of rice NCED knockout mutants.

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Heterologous expression of rice 9-cis-epoxycarotenoid dioxygenase 4 (OsNCED4) in Arabidopsis confers sugar oversensitivity and drought tolerance

Hwang et al. Bot Stud Heterologous expression of rice 9-cis-epoxycarotenoid dioxygenase 4 (OsNCED4) in Arabidopsis confers sugar oversensitivity and drought tolerance San‑Gwang Hwang 0 ChiaY‑un Lee 2 Ching‑Shan Tseng 1 0 Department of Horticulture, National Chung Hsing University , 145 Xingda Road, South District, Taichung 40227, Taiwan, ROC 1 Taiwan Agricultural Research Institute, Council of Agriculture , Executive Yuan, No.189, Zhongzheng Road, Wufeng District, Taichung 41362, Taiwan, ROC 2 Institute of Plant and Microbial Biology, Academia Sinica , Taipei 11529, Taiwan, ROC Background: The 9‑ cis‑ epoxycarotenoid dioxygenases OsNCED4 was cloned from rice in conjunction with OsNCED 1‑ 3 and 5, of which 3 has been shown to function in ABA biosynthesis and alteration of leaf morphology. In higher plants, NCEDs have been shown to be key enzymes controlling ABA biosynthesis and belong to a differentially expressed gene family. Aside from OsNCED3, it remains largely unknown if other OsNCED genes are involved in ABA biosynthesis in rice. Thus, transgenic Arabidopsis plants overexpressing OsNCED4 were generated in the 129B08/nced3 mutant background to explore OsNCED4 function in ABA biosynthesis. Results: Heterologous expression of OsNCED4 in Arabidopsis increased ABA levels and altered plant size and leaf shape, delayed seed germination, caused sugar oversensitivity in post‑ germination growth, and enhanced tolerance to drought. The native OsNCED3 and OsNCED4 promoters were expressed in an overlapping pattern in rice seeds and young seedlings, suggesting possible functional redundancy between OsNCED3 and OsNCED4. At the one‑ leaf stage, similar regulation of OsNCED3 and OsNCED4 gene expression in roots or leaves in response to moderate salt stress (150 mM NaCl) was observed. Conclusion: Like OsNCED3, OsNCED4 is functionally active in ABA biosynthesis in rice. OsNCED3 and OsNCED4 might play redundant roles in controlling ABA biosynthesis in rice, as suggested by GUS staining assay, but this should be further analyzed through complementation of rice NCED knockout mutants. Abscisic acid; 9‑ cis‑ epoxycarotenoid dioxygenase; Seed germination; Post‑ germination growth arrest; Drought tolerance Background The functions of abscisic acid (ABA) in regulation of seed dormancy, seedling growth and development, stomatal closure, and stress tolerance is well studied (Jiang and Yu 2009; Zhu et  al. 2009, 2011; Waterland et  al. 2010; Gao et al. 2011; Bauer et al. 2013; Merilo et al. 2015) . Connections among ABA, molecular signaling components, and nutrition have been identified. For example, Jiang and Yu (2009) suggested that the transcription factor AtWRKY2 regulates seed germination and post-germination developmental arrest in Arabidopsis via its response to ABA. At the same year, Zhu et al. (2009) reported that the glucose-induced germination delay in rice seeds is due to the prevention of ABA degradation, rather than an increase in ABA biosynthesis. Zhu et al. (2011) further elaborated that downregulation of CYP707A2 expression, a gene encoding an ABA 8′-hydroxylase involved in ABA catabolism, and the subsequent reduction in ABA degradation, are closely associated with the delay of seed germination and seedling growth in Arabidopsis. More recently, Gao et  al. (2011) characterized the protein function of AtCPR5 and revealed that seed germination and early seedling growth are independently regulated through the ABA and lipoxygenase (LOX) pathways. ABA regulates numerous physiological responses in addition to seed germination and seedling growth. Previous reports have shown that ABA synthesis in guard cells is essential and sufficient for stomatal closure in response to declined relative humidity (Bauer et  al. 2013; Merilo et  al. 2015) . Waterland et  al. (2010) demonstrated the involvement of ABA in drought tolerance. The roles of ABA and ABA signaling in plant abiotic stress responses, including drought tolerance, have been recently reviewed and discussed (Sah et al. 2016; Vishwakarma et al. 2017) . In higher plants, 9-cis-epoxycarotenoid dioxygenases (NCEDs) are thought to be the key enzymes controlling ABA biosynthesis and stress tolerance (Iuchi et  al. 2001; Sun et al. 2012; Vishwakarma et al. 2017) . To date, five rice NCED genes have been reported (Oliver et  al. 2007; Welsch et  al. 2008; Zhu et  al. 2009) . However, only OsNCED3 (GenBank Accession No. AY838899) was characterized as functionally active in ABA biosynthesis (Hwang et  al. 2010) . The biological functions of the other four rice NCED genes, OsNCED1 (GenBank Accession No. AY838897), OsNCED2 (GenBank Accession No. AY838898), OsNCED4 (GenBank Accession No. AY838900) and OsNCED5 (GenBank Accession No. AY838901) have yet to be deciphered. It remains unknown whether rice OsNCED4 gene is involved in ABA biosynthesis or ABA-regulated physiological processes. Previous studies of the NCED gene family in Arabidopsis revealed that diffe (...truncated)


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San-Gwang Hwang, Chia-Yun Lee, Ching-Shan Tseng. Heterologous expression of rice 9-cis-epoxycarotenoid dioxygenase 4 (OsNCED4) in Arabidopsis confers sugar oversensitivity and drought tolerance, Botanical Studies, 2018, pp. 2, Volume 59, Issue 1, DOI: 10.1186/s40529-018-0219-9