A VIT‐like transporter facilitates iron transport into nodule symbiosomes for nitrogen fixation in soybean

Abstract

Effective legume-rhizobia symbiosis depends on efficient nutrient exchange. Rhizobia need to synthesize iron-containing proteins for symbiotic nitrogen fixation (SNF) in nodules, which depends on host plant-mediated iron uptake into the symbiosome.We functionally investigated a pair of vacuolar iron transporter like (VTL) genes, GmVTL1a/b, in soybean (Glycine max) and evaluated their contributions to SNF, including investigations of gene expression patterns, subcellular localization, and mutant phenotypes. Though both GmVTL1a/b genes were specifically expressed in the fixation zone of the nodule, GmVTL1a was the lone member to be localized at the tonoplast of tobacco protoplasts, and shown to facilitate ferrous iron transport in yeast. GmVTL1a targets the symbiosome in infected cells, as verified by in situ immunostaining. Two vtl1 knockout mutants had lower iron concentrations in nodule cell sap and peribacteroid units than in wild-type plants, suggesting that GmVTL1 knockout inhibited iron import into symbiosomes. Furthermore, GmVTL1 knockout minimally affected soybean growth under nonsymbiotic conditions, but dramatically impaired nodule development and SNF activity under nitrogen-limited and rhizobia-inoculation conditions, which eventually led to growth retardation.Taken together, these results demonstrate that GmVTL1a is indispensable for SNF in nodules as a transporter of ferrous iron from the infected root cell cytosol to the symbiosome.

GmVTL1a is an iron transporter on the symbiosome membrane of soybean with an important role in nitrogen fixation

“…In a very recent paper, Liu et al . (2020) reported similar results for GmVTL1a. Our proteomic results for GmVTL1b suggest that the identification of GmVTL1b in the ER by Lui et al .…”

“…These results link the mutations that abolish nitrogen fixation in SEN1 to the function as an iron transporter. The similar phenotype for sen1 and vtl1‐1 mutants in terms of nodule numbers, appearance, reduction in nitrogen fixation and in nodule iron content (this study, Suganuma et al ., 2003; Hakoyama et al ., 2012; Lui et al ., 2020) and the ability of GmVTL1a to complement the sen1 mutant suggests that LjSEN1 and GmVTL1a play the same role. Furthermore, this suggests GmVTL1a is important for proper functioning of the symbiosis.…”

“…The reduction of iron in the cytoplasm of the infected cells is shown through Perls/DAB staining of sen1‐1 (this study) and vtl1‐1 nodules (Liu et al ., 2020). These findings combined with reduced iron content in vtl1‐1 nodules (Liu et al ., 2020) suggest feedback regulation reduces transport of iron into infected cells or nodules when transport into the symbiosome is blocked.…”

Vacuolar Iron Transporter (Like) proteins: Regulators of cellular iron accumulation in plants

“…Till recently, soybean DMT1 (Divalent Metal Transporter 1) has been shown to be the only Fe transporter to nodules (Kaiser et al, 2003). However, recently there have been at least three independent reports that demonstrate the role of VTL genes in Fe transport in symbiotic nodules in legume crops (Brear et al, 2020; Liu et al, 2020; Walton et al, 2020).…”

Medicago truncatula Ferroportin2 mediates iron import into nodule symbiosomes

“…Mutation of this gene in different legumes results in loss of nitrogen fixation capabilities, defective nodule development, and altered iron homeostasis (Hakoyama et al ., 2012; Walton et al ., 2019; Brear et al ., 2020; Liu et al ., 2020). Supporting this tentative role in the leaky fpn2 phenotype are the recent results on its subcellular localization that show that SEN1 (renamed GmVTL1 in soybean, and MtVTL8 in M. truncatula ) is likely located in the symbiosomes (Walton et al ., 2019; Brear et al ., 2020; Liu et al ., 2020). Therefore, M. truncatula has very likely two active symbiosome iron uptake systems which are not functionally redundant, since mutating either has a strong impact on nodulation and nitrogen fixation.…”