Functional analysis of the novel mycorrhiza‐specific phosphate transporter AsPT1 and PHT1 family from Astragalus sinicus during the arbuscular mycorrhizal symbiosis

Abstract

SummaryArbuscular mycorrhizas contribute significantly to inorganic phosphate (Pi) uptake in plants. Gene networks involved in the regulation and function of the Pht1 family transporters in legume species during AM symbiosis are not fully understood.In order to characterize the six distinct members of Pht1 transporters in mycorrhizal Astragalus sinicus, we combined cellular localization, heterologous functional expression in yeast with expression/subcellular localization studies and reverse genetics approaches in planta. Pht1;1 and Pht1;4 silenced lines were generated to uncover the role of the newly discovered dependence of the AM symbiosis on another phosphate transporter AsPT1 besides AsPT4.These Pht1 transporters are triggered in Pi-starved mycorrhizal roots. AsPT1 and AsPT4 were localized in arbuscule-containing cells of the cortex. The analysis of promoter sequences revealed conserved motifs in both AsPT1 and AsPT4. AsPT1 overexpression showed higher mycorrhization levels than controls for parameters analysed, including abundance of arbuscules. By contrast, knockdown of AsPT1 by RNA interference led to degenerating or dead arbuscule phenotypes identical to that of AsPT4 silencing lines. AsPT4 but not AsPT1 is required for symbiotic Pi uptake.These results suggest that both, AsPT1 and AsPT4, are required for the AM symbiosis, most importantly, AsPT1 may serve as a novel symbiotic transporter for AM development.

Arbuscular Mycorrhizal Symbiosis Requires a Phosphate Transceptor in the Gigaspora margarita Fungal Symbiont

“…The dynamics of arbuscule level in roots was confirmed by the expression of AM-specific Pi transporter gene AsPT4, which is particularly expressed in arbusculated cells and therefore an indicator of the function of AM symbiosis ( Xie et al, 2013). It was found that the expression profiles of GigmPT were similar to that of AsPT4 over time ( Figure 3B).…”

“…The subcellular localization of GigmPT in yeast was examined with a C-terminal fusion of GigmPT to GFP in the yeast strain as described by Xie et al (2013). The image capture was performed using a fluorescence microscope (Olympus BX51, Japan).…”

“…To detect the activity of the GigmPT gene in HIGS mutants, we examined expression levels of the fungal GigmPT, GigmPT1, and the AM-specific PT genes AsPT1 and AsPT4 ( Xie et al, 2013) by qRT-PCR in mycorrhizal roots. Transcription of GigmPT was significantly reduced in the positive RNAi roots ( Figure 6J and Supplemental Figure 9D).…”

“…G. margarita was employed to inoculate the roots of Astragalus sinicus L. as described elsewhere ( Xie et al, 2013). Spores of G. margarita were produced in pot system on A. sinicus and collected by wet sieving, after which the spores were soaked in 2% Chloramine T (Sigma-Aldrich, St. Louis, MO, USA) for 15 min, washed at least three times in sterile water, and immersed in a mixture of antibiotic solution containing 200 mg l−1 streptomycin and 100 mg l−1 gentamycin (Sigma-Aldrich) for 10 min.…”

Presidential address: recent advance of mycorrhizal research in China

“…Liu et al (2015) found that drought stress upregulated the level of R. intraradices and soybean mitogen-activated protein kinase (MAPK) transcripts in mycorrhizal soybean roots, indicating the possibility of a molecular dialogue between AM fungus and host plant and suggesting that they might cooperate to improve soybean’s resistance to drought stress. Concurrently, in terms of nutrient absorption, some studies investigated the mycorrhiza-regulated phosphate transporters in host plants Astragalus sinicus (Xie et al 2013), Oryza sativa (Sun et al 2012), P. tabulaeformis (Zheng et al 2016a), pepper and tobacco (Chen et al 2007), and reciprocal transportation of carbon and phosphorus amongst plants ( Zea mays and Medicago sativa ), Rhizophagus irregularis and phosphate-solubilising bacterium (Zhang et al 2016; Wang et al 2016a). Wang et al (2014) demonstrated that H + -ATPase played a key role in energising the periarbuscular membrane, thereby facilitating nutrient exchange in arbusculated rice and Medicago truncatula cells.…”