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Acta Agron Sin ›› 2013, Vol. 39 ›› Issue (11): 1952-1961.doi: 10.3724/SP.J.1006.2013.01952

• CROP GENETICS & BREEDING · GERMPLASM RESOURCES · MOLECULAR GENETICS • Previous Articles     Next Articles

Characteristic and Function Analysis of a Copper Ion Binding Protein, AtBCB Interacting with G Protein α Subunit GPA1 in Arabidopsis thaliana

ZHANG Xiao-Hong1,XU Peng-Bo1,2,GUO Meng-Meng1,2,XU Zhao-Shi2,LI Lian-Cheng2,CHEN Ming2,*,MA You-Zhi2   

  1. 1 College of Life Sciences, Northwest A&F University, Yangling 712100, China; 2 Institute of Crop Sciences, Chinese Academy of Agricultural Sciences / National Key Facility for Crop Gene Resources and Genetic Improvement, Key Laboratory of Biology and Genetic Improvement of Triticeae Crops, Ministry of Agriculture, Beijing 100081
  • Received:2013-04-07 Revised:2013-06-09 Online:2013-11-12 Published:2013-08-01
  • Contact: 陈明, E-mail: chenming02@caas.cn; Tel: 010-82108750

Abstract:

Heterotrimeric G protein, including three subunits of α, β, and γ, is involved in a lot of signaling pathways in plants. It receives extracellular signals via G-protein coupled receptor (GPCR) and transmits them to the downstream effectors by the three subunits. Till now, the downstream effectors and signaling pathway related to G-protein complexes have been rarely reported. Furthermore, identifying more novel G protein effectors would be helpful to elucidate signaling pathway associated with the G protein complex. In order to find some novel effectors, G protein α subunit (GPA1) was used as bait to screen interaction protein in Arabidopsis by the split-ubiquitin screening system in this study. One of the GPA1-interacting proteins was identified as copper ion binding protein,AtBCB. The interaction betweenGPA1 and AtBCB was verified on cell membrane by BiFC (bimolecular fluorescence complementation). The expressions of GPA1 and AtBCB were confirmed to be induced by aluminium stress. To study the function of the two genes, we treated Arabidopsis mutant gpa1-4 and bcb, in which GPA1 and AtBCB were knocked out, with 100 µmol L–1 Al3+, respectively, and then measured MDA (malonaldehyde) content in roots. The results showed that MDA content in both mutant and WT (wild type) under normal condition was no significant difference, but when exposured to 100 µmol L–1 Al3+ the content in gpa1-4 was lower than that in WT (P<0.05), and it in bcb was higher than that in WT (P<0.01). Furthermore, the expression patterns of three responsive genes ofALMT1, ALS1 and ALS3 to aluminum toxicity were analyzed by Real-time PCR. The results showed that no matter the condition with or without aluminum stress, the expression of ALS1 and ALS3 appeared no significant difference in the mutants and WT. However, in the treatment with less than 100 µmol L–1 Al the expression level of ALMT1 ingpa1-4 was higher than that in WT, and the expression level of ALMT1 inbcbwas lower than that in WT. In short, GPA1 and AtBCB directly interact in the cell membrane, and regulate the expression of the downstream gene of ALMT1. In the tolerance process to aluminum stress in plants, GPA1 plays negative role but AtBCB has positive effect.

Key words: G protein, Split-ubiquitin system, Protein interaction, Aluminium ion stress, BiFC

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