Accumulation of reactive oxygen species in arbuscular mycorrhizal roots

Abstract

We investigated the accumulation of reactive oxygen species (ROS) in arbuscular mycorrhizal (AM) roots from Medicago truncatula, Zea mays and Nicotiana tabacum using three independent staining techniques. Colonized root cortical cells and the symbiotic fungal partner were observed to be involved in the production of ROS. Extraradical hyphae and spores from Glomus intraradices accumulated small levels of ROS within their cell wall and produced ROS within the cytoplasm in response to stress. Within AM roots, we observed a certain correlation of arbuscular senescence and H2O2 accumulation after staining by diaminobenzidine (DAB) and a more general accumulation of ROS close to fungal structures when using dihydrorhodamine 123 (DHR 123) for staining. According to electron microscopical analysis of AM roots from Z. mays after staining by CeCl3, intracellular accumulation of H2O2 was observed in the plant cytoplasm close to intact and collapsing fungal structures, whereas intercellular H2O2 was located on the surface of fungal hyphae. These characteristics of ROS accumulation in AM roots suggest similarities to ROS accumulation during the senescence of legume root nodules.

Reactive oxygen species and plant resistance to fungal pathogens

“…The cytochemical staining using cerium (III) chloride (CeCl 3 ) is used for a subcellular localisation of H 2 O 2 . The reaction between CeCl 3 and an excess of H 2 O 2 generates electron-dense deposits of cerium perhydroxides that can be observed using transmission electron microscopy (Bestwick et al, 1998;Fester and Hause, 2005;Lherminier et al, 2009;Simon et al, 2013;Xia et al, 2009).…”

“…They consist in non-fluorescent molecules that become fluorescent when oxidised by ROS, and the emitted fluorescence can be observed by fluorimetry and/or by fluorescent microscopy, an advantage of such probes (Benikhlef et al, 2013;Bulgakov et al, 2012;Fester and Hause, 2005;Guo et al, 2010;Kolla et al, 2007;L'Haridon et al, 2011;Li et al, 2007;Liu et al, 2010;Ma et al, 2013;Peleg-Grossman et al, 2012;Plancot et al, 2013;Tada et al, 2004;Wen et al, 2008;Ye et al, 2013). Luminol or luminol analogues are sensitive chemiluminescent probes used to quantify a relative intensity of ROS by counting the emitted light with a luminometer, CDD camera or a scintillation counter (Dubreuil-Maurizi et al, 2010;Flury et al, 2013;Kunz et al, 2006;L'Haridon et al, 2011;Mersmann et al, 2010).…”

Reactive oxygen species generation and signaling in plants

“…128 ROS might have different functions during various steps of the beneficial symbiosis, however it is generally excepted that ROS accumulates during early phases of the interaction when the beneficial symbiosis is not yet stable and declines thereafter. 129 Whether this is caused by a general reduction in host defense processes once the two symbionts have been recognized as friends, or caused by an active repression from the microbe, is not known yet. Beneficial fungi or microbe are also known to protect plants against pathogens, e.g.…”

Characterization of a CuZn superoxide dismutase gene in the arbuscular mycorrhizal fungus Glomus intraradices

“…These structures, called arbuscules, are thought to be the main sites of nutrient and signal exchange between the two partners (Parniske 2008). An accumulation of H2O2 and catalase and peroxidase transcripts has been observed in arbuscule-containing root cells of different plant species (Blee and Anderson 2000; Lambais et al 2003;Fester and Hause 2005). Far less is known about the antioxidant responses on the fungal partner.…”

“…The development of AM in vitro cultures, that is the co-culture of transformed hairy roots and AM fungi, has allowed the investigation of soil-free extraradical hyphae and spores (Fortin et al 2002). By using this experimental system, it has been shown that environmental stresses, such as exposure to Cu or mechanical damage, promote ROS formation in the extraradical mycelium of Glomus intraradices (Fester and Hause 2005;Benabdellah et al 2009). To cope with this oxidative stress, AM fungi must activate their antioxidant defences.…”