Elsevier

Journal of Proteomics

Volume 74, Issue 8, 12 August 2011, Pages 1338-1350
Journal of Proteomics

Proteomic analysis as a tool for investigating arsenic stress in Pteris vittata roots colonized or not by arbuscular mycorrhizal symbiosis

https://doi.org/10.1016/j.jprot.2011.03.027Get rights and content

Abstract

Pteris vittata can tolerate very high soil arsenic concentration and rapidly accumulates the metalloid in its fronds. However, its tolerance to arsenic has not been completely explored. Arbuscular mycorrhizal (AM) fungi colonize the root of most terrestrial plants, including ferns. Mycorrhizae are known to affect plant responses in many ways: improving plant nutrition, promoting plant tolerance or resistance to pathogens, drought, salinity and heavy metal stresses. It has been observed that plants growing on arsenic polluted soils are usually mycorrhizal and that AM fungi enhance arsenic tolerance in a number of plant species. The aim of the present work was to study the effects of the AM fungus Glomus mosseae on P. vittata plants treated with arsenic using a proteomic approach. Image analysis showed that 37 spots were differently affected (21 identified). Arsenic treatment affected the expression of 14 spots (12 up-regulated and 2 down-regulated), while in presence of G. mosseae modulated 3 spots (1 up-regulated and 2 down-regulated). G. mosseae, in absence of arsenic, modulated 17 spots (13 up-regulated and 4 down-regulated). Arsenic stress was observed even in an arsenic tolerant plant as P. vittata and a protective effect of AM symbiosis toward arsenic stress was observed.

Introduction

Arsenic (As) is a metalloid occurring in the earth crust and naturally released in the environment as consequence of erosion, volcanic emissions, etc. Elevated arsenic concentrations in soils have been found also in areas impacted by mining and smelting industries, by coal burning [1] and by the use of arsenic containing agrochemicals [2]. Arsenic is highly toxic to most biological systems and carcinogenic for man [3] and can diffuse in soils and groundwater, so entering the food chain through drinking water and contaminated vegetables [4].

Plants have different sensitivity to arsenic, with legumes known to be highly sensitive [5]. Pteris vittata L. (Chinese brake fern), the first known arsenic-hyperaccumulating plant, is able to remove large amounts of arsenic from soil [6]. The fern shows interesting growth characteristics, including a large biomass, extensive root system, high growth rate and perennial habit. Typical of hyperaccumulators, arsenic in P. vittata is mostly concentrated in the fronds [6], [7] even if arsenic concentration in roots can be high (near 100 mg/kg). This arsenic concentration can be considered very toxic for the majority of plant species but not for P. vittata that can tolerate up to 10,000 mg/kg [8]. By contrast, as reported in Kabata-Pendias 2001 [9], in plant tissues, non toxic arsenic concentration range between 1 and 1.7 mg/kg (ppm) and toxic concentration range between 5 and 20 mg/kg [9]. However, in spite of the very high capability of P. vittata to tolerate arsenic, this tolerance is not complete, as recently shown by proteome analyses of the fronds [7].

Roots are the sole access point to below-ground trace elements and as such they play a vital role in hyperaccumulation. Their role as an effective trace element scavenger is achieved through interactions in the rhizosphere with bacteria and fungi, among which arbuscular mycorrhizal (AM) fungi, a monophyletic group of soil fungi in the Glomeromycota phylum [10] that colonize most terrestrial plants [11], [12]. This symbiosis not only plays a central role in soil nutrient uptake but also improves plant tolerance to biotic and abiotic stresses [13], [14], [15].

Proteomic analysis has been used extensively to investigate the protein expression pattern under several abiotic stresses. The expression pattern of maize root proteins in response to arsenic stress has been described for the first time by Requejo and Tena (2005) [16] and the arsenic-induced differentially expressed proteins in rice roots have been described [17]. Some information exists on protein expression profile induced by AM fungi in plant roots [18], [19], [20] and a number of reports show the influence of AM fungi and other soil microorganisms in protection against protein modification induced by heavy metals [7], [21], [22]. Nevertheless, as far as we know, no data are available on AM fungi effects on arsenic absorption at root level in the fern P. vittata and on physiological changes and protein expression profile modifications induced by these fungi. Therefore, the aim of this study was to investigate the effects induced in the protein expression profile of P. vittata roots by the AM fungus Glomus mosseae in presence and in absence of arsenic contamination.

Section snippets

Experimental design and plant material

The plants used in these experiments were obtained from spores collected at the Botanical Garden of Genoa (Italy) and grown in controlled conditions (16/8 h light/dark photoperiod, 150 μmol/m2 s light irradiance and 24 °C/20 °C light/dark thermoperiod). At least twelve replicates per each treatment were analyzed.

The experimental design included four different theses: control plants (C) (plants grown on quartz sand fed only with Long Ashton nutrient solution [23] containing 32 μM phosphate (added as

Effect of arsenic and AM fungi on morphological parameters

Ferns were treated weekly (after a month of acclimation), for sixty days, with 25 mg/l of arsenic, a non-lethal dose for P. vittata, in order to evaluate the effect of chronic arsenic contamination. As plants showed a statistically significant higher biomass both in roots and in fronds compared to C plants (Fig. 1). Gm plants showed a statistically significant higher dry weight compared to C plants both in frond but especially in roots that showed a dry biomass about three times higher compared

Arsenic tolerance in ferns inoculated or not with G. mosseae

Our experimental design has been planned in order to improve G. mosseae colonization and relieve transplanting stress, the acclimation time has been prolonged to one month, differing from previous data on P. vittata frond proteome [7]. So M% and A% increased in GmAs plants, in accordance to the process referred as priming [31]. The higher colonization percentage leads to a statistical significant increase of dry weight of Gm roots and fronds.

Arsenic content evaluation confirmed that P. vittata

Acknowledgments

This research was funded by the Italian Minister for Education and Scientific Research, PRIN 2007 program- prot. 2007PKFAAT, Project title: mechanisms of response to arsenic and cadmium in model plants: from molecular level to in field investigation.

The authors have declared no conflict of interest.

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