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Plant and Cell Physiology Advance Access published online on November 19, 2008
Plant and Cell Physiology, doi:10.1093/pcp/pcn178
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© The Author 2008. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For Permissions, please e-mail: journals.permissions@oxfordjournals.org

Nitric oxide regulates shikonin formation in suspension cultured Onosma paniculatum cells

Shu-Jing Wu*,1, Jin-Liang Qi*,1, Wen-Ju Zhang*,1, Shao-Hua Liu1, Feng-Hui Xiao2, Ming-Sheng Zhang1, Guo-Hua Xu1, Wei-Guo Zhao1, Ming-Wang Shi1, Yan-Jun Pang1, Heng-Guan Shen1 and Yong-Hua Yang**,1

1Institute of Plant Molecular Biology, State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing 210093, P. R. China,
2Department of Biotechnology, Yunnan Agricultural University, Kunming 650201, P. R. China

**Corresponding author: Prof. Yong-Hua Yang. Institute of Plant Molecular Biology, State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing 210093, P. R. China. TEL: 86-25-83686305, FAX: 86-25-83686305, E-mail: YangYH{at}nju.edu.cn


  Abstract

Endogenously occurred nitric oxide (NO) is involved in the regulation of shikonin formation in Onosma paniculatum cells. NO generated after cells were inoculated into shikonin production medium, reaching the highest level after 2 d of culture, which is 16 times of that at the beginning, and maintaining at a high level within 6 days. A nitric oxide synthase (NOS) inhibitor, N{omega}-Nitro-L-arginine (L-NNA), and a nitrate reductase (NR) inhibitor, sodium azide (SoA), in consistent with their inhibition on NO biosynthesis, decreased shikonin formation significantly. This reduction could be alleviated or even abolished by exogenous NO supplied by sodium nitroprusside (SNP), suggesting that the inhibition of NO biosynthesis resulted in decreased shikonin formation. However, when endogenous NO biosynthesis was up-regulated by the elicitor from Rhizoctonia cerealis, shikonin production was enhanced furthermore, showing a dependence on the elicitor-induced NO outburst. Real-time PCR analysis showed that NO could significantly up-regulate the expression of PAL, PGT and HMGR, which encode key enzymes involved in shikonin biosynthesis. These results demonstrated that NO plays a critical role in shikonin formation in O. paniculatum cells.

Keywords: Electron paramagnetic resonance (EPR) - Fungal elicitor - Nitric oxide - Onosma paniculatum - Shikonin

*These authors contributed equally to this work

(Received August 19, 2008; Accepted November 13, 2008)
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