GintAMT3 – a Low-Affinity Ammonium Transporter of the Arbuscular Mycorrhizal Rhizophagus irregularis

Abstract

Nutrient acquisition and transfer are essential steps in the arbuscular mycorrhizal (AM) symbiosis, which is formed by the majority of land plants. Mineral nutrients are taken up by AM fungi from the soil and transferred to the plant partner. Within the cortical plant root cells the fungal hyphae form tree-like structures (arbuscules) where the nutrients are released to the plant-fungal interface, i.e., to the periarbuscular space, before being taken up by the plant. In exchange, the AM fungi receive carbohydrates from the plant host. Besides the well-studied uptake of phosphorus (P), the uptake and transfer of nitrogen (N) plays a crucial role in this mutualistic interaction. In the AM fungus Rhizophagus irregularis (formerly called Glomus intraradices), two ammonium transporters (AMT) were previously described, namely GintAMT1 and GintAMT2. Here, we report the identification and characterization of a newly identified R. irregularis AMT, GintAMT3. Phylogenetic analyses revealed high sequence similarity to previously identified AM fungal AMTs and a clear separation from other fungal AMTs. Topological analysis indicated GintAMT3 to be a membrane bound pore forming protein, and GFP tagging showed it to be highly expressed in the intraradical mycelium of a fully established AM symbiosis. Expression of GintAMT3 in yeast successfully complemented the yeast AMT triple deletion mutant (MATa ura3 mep1Δ mep2Δ::LEU2 mep3Δ::KanMX2). GintAMT3 is characterized as a low affinity transport system with an apparent Km of 1.8 mM and a Vmax of 240 nmol-1 min-1 108 cells-1, which is regulated by substrate concentration and carbon supply.

Fungal and plant gene expression in the Tulasnella calospora–Serapias vomeracea symbiosis provides clues about nitrogen pathways in orchid mycorrhizas

“…Nitrate transporters and genes responsible for nitrate utilization have been identified mainly in AM fungi (Kaldorf et al, 1998;Tian et al, 2010;Koegel et al, 2015), and nitrophilous ECM fungi (Plassard et al, 2000;Jargeat et al, 2003;Montanini et al, 2006). Genes coding for high-and low-affinity ammonium transporters have been identified and characterized in both AM (L opez-Pedrosa et al, 2006;P erez-Tienda et al, 2011;Calabrese et al, 2016) and ECM fungi (Javelle et al, 2001;Montanini et al, 2002;Willmann et al, 2007). Amino acid permeases and peptide transporters, which allow for organic N uptake, have been identified and characterized mainly in ECM fungi (Nehls et al, 1999;Wipf et al, 2002;Guidot et al, 2005;Benjdia et al, 2006;Shah et al, 2013) but also in AM fungi (Cappellazzo et al, 2008;Belmondo et al, 2014), although the ability of AM fungi to utilize organic N sources is considered to be relatively low.…”

Identification of arbuscular mycorrhiza-inducible Nitrate Transporter 1/Peptide Transporter Family (NPF) genes in rice

“…In the AMF Rhizophagus intraradices (formerly called Glomus intraradices), that is found in rice roots in paddy fields and is used in most AM-rice studies (e.g. Paszkowski et al 2002;Güimil et al 2005;Glassop et al 2007;Gutjahr et al 2008), the ammonium transporters AMT1, AMT2 and AMT3 are expressed in the intraradical mycelium (Lopez-Pedrosa et al 2006;Perez-Tienda et al 2011;Calabrese et al 2016), suggesting a participation in retrieval of NH 4 + leaking from the fungus during fungal metabolism at the symbiotic interface. Alternatively, ammonium released by the AM at this interface may be taken up by the plant.…”

Phylogenetic, structural, and functional characterization of AMT3;1, an ammonium transporter induced by mycorrhization among model grasses

“…Three high-affinity AMTs were identified in the AM fungus Rhizophagus irregularis: GintAMT1 (López-Pedrosa et al 2006), GintAMT2 (Pérez-Tienda et al 2011, and GintAMT3 (Calabrese et al 2016). GintAMT1 is expressed in the extraradical mycelium, suggesting a role in NH 4 + acquisition from the soil, while GintAMT2 might be involved in retrieving leaked NH 4 + .…”