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JXB Advance Access originally published online on April 30, 2009
Journal of Experimental Botany 2009 60(9):2613-2620; doi:10.1093/jxb/erp104
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© 2009 The Author(s).
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. This paper is available online free of all access charges (see http://jxb.oxfordjournals.org/open_access.html for further details)

RESEARCH PAPER

Characterization of a canola C2 domain gene that interacts with PG, an effector of the necrotrophic fungus Sclerotinia sclerotiorum

Xinyu Wang1,2,3 *,{dagger}, Qian Li1 *, Xiaowei Niu2, Haiyan Chen1, Langlai Xu1 and Cunkou Qi3,{dagger}

1College of Life Sciences, Nanjing Agricultural University, Nanjing 210095, China
2The State Key Laboratory of Crop Genetics & Germplasm enhancement, Nanjing Agricultural University, Nanjing 210095, China
3Academy of Jiangsu Agricultural Sciences, Nanjing 210014, China

{dagger} To whom correspondence should be addressed. E-mail: xywang{at}njau.edu.cn, qck{at}jaas.ac.cn

Sspg1d, one of endopolygalacturonases, is an important fungal effector secreted by the necrotrophic fungus Sclerotinia sclerotiorum during early infection. Using sspg1d as bait, a small C2 domain protein (designated as IPG-1) was identified by yeast two-hybrid screening of a canola cDNA library. Deletion analysis confirmed that the C-terminus of IPG-1 is responsible for its interaction with sspg1d in the yeast two-hybrid assay. The sspg1d/IPG-1 interaction was further confirmed in plant cells by a biomolecular fluorescence complementation (BiFC) assay. A transient expression assay showed that the IPG-1–GFP fusion protein was targeted to the plasma membrane and nucleus in onion epidermal cells. Following treatment with a Ca2+ ionophore, it was distributed throughout the cytosol. Real-time PCR assay demonstrated that IPG-1 was highly induced by Sclerotinia sclerotiorum in canola leaves and stems. Southern blot analysis indicated the presence of about five homologues of IPG-1 in the canola genome. Two additional members of the IPG-1gene family were isolated by RT-PCR. Their sequence similarity with IPG-1 is as high as 95%. However, they did not interact with sspg1d in the yeast two-hybrid assay. Possible roles of IPG-1 and its association with sspg1d in the defence signalling pathway were discussed.

Key words: BiFC, C2 domain, endo-PG, PG, Sclerotinia sclerotiorum

* These authors contributed equally to this work.

Received 19 January 2009; Revised 11 March 2009 Accepted 11 March 2009


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