ITRAQ analysis reveals mechanisms of growth defects due to excess zinc in Arabidopsis.

Yoichiro Fukao,Nahoko Nagasaki,Masayuki Fujiwara,Rie Kurata,Masayoshi Maeshima,Rie Tomioka,Yuka Nishimori,Ali Ferjani

doi:10.1104/pp.110.169730

Yoichiro Fukao, Nahoko Nagasaki + Show 6 more

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https://doi.org/10.1104/pp.110.169730

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Abstract

The micronutrient zinc is essential for all living organisms, but it is toxic at high concentrations. Here, to understand the effects of excess zinc on plant cells, we performed an iTRAQ (for isobaric tags for relative and absolute quantification)-based quantitative proteomics approach to analyze microsomal proteins from Arabidopsis (Arabidopsis thaliana) roots. Our approach was sensitive enough to identify 521 proteins, including several membrane proteins. Among them, IRT1, an iron and zinc transporter, and FRO2, a ferric-chelate reductase, increased greatly in response to excess zinc. The expression of these two genes has been previously reported to increase under iron-deficient conditions. Indeed, the concentration of iron was significantly decreased in roots and shoots under excess zinc. Also, seven subunits of the vacuolar H(+)-ATPase (V-ATPase), a proton pump on the tonoplast and endosome, were identified, and three of them decreased significantly in response to excess zinc. In addition, excess zinc in the wild type decreased V-ATPase activity and length of roots and cells to levels comparable to those of the untreated de-etiolated3-1 mutant, which bears a mutation in V-ATPase subunit C. Interestingly, excess zinc led to the formation of branched and abnormally shaped root hairs, a phenotype that correlates with decreased levels of proteins of several root hair-defective mutants. Our results point out mechanisms of growth defects caused by excess zinc in which cross talk between iron and zinc homeostasis and V-ATPase activity might play a central role.

Highlights

The micronutrient zinc is essential for all living organisms, but it is toxic at high concentrations
Microsomal proteins were prepared from roots grown for 10 d on Murashige and Skoog (MS) medium already containing 30 mM ZnSO4 or MS medium exogenously supplemented with 300 mM ZnSO4
We summarized results using less than 5% false discovery rate (FDR) in each iTRAQ analysis (FDR = 4.06, 3.13, or 3.62; Elias and Gygi, 2007)

Summary

Introduction

The micronutrient zinc is essential for all living organisms, but it is toxic at high concentrations. It has been proposed that such growth defects might be a secondary effect of excess zinc caused by deficiency of other essential ions, such as iron or magnesium, which have ionic radii similar to zinc (Marschner, 1995). As a strategy to avoid the toxicity of several ions, plant cells take advantage of such proton gradients to sequester the ions inside vacuoles (Dietz et al, 2001) During this process, the vacuolar H+-ATPase (V-ATPase), which is a heteromultimeric proton pump, together with H+-pyrophosphatase (V-PPase), plays a central role in vacuole acidification, energizing the active transport of ions across the tonoplast and coordinately regulating turgor pressure and cell elongation (Maeshima, 2000; Sze et al, 2002). The V-ATPase plays an important role in the trans-Golgi network (TGN) for cell expansion, which is involved in the synthesis and trafficking of cell wall components, such as cellulose (Brux et al, 2008)

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