Seminal and nodal roots of barley differ in anatomy, proteome and nitrate uptake capacity

The root system of barley plants is composed of embryogenic, seminal roots as well as lateral and nodal roots that are formed postembryonically from seminal roots and from the basal part of shoots, respectively. Due to their distinct developmental origin, seminal and nodal roots may differ in function during plant development; however, a clear comparison between these two root types has not yet been undertaken. In this study, anatomical, proteomic and physiological traits were compared between seminal and nodal roots of similar developmental stages. Nodal roots have larger diameter, larger metaxylem area and a larger number of metaxylem vessels than seminal roots. Proteome profiling uncovered a set of root-type-specific proteins, including proteins related to the cell wall and cytoskeleton organization, which could potentially be implicated with differential metaxylem development. We also found that nodal roots have higher levels of auxin, which is known to trigger metaxylem development. At millimolar nitrate supply, nodal roots had approximately 2-fold higher nitrate uptake and root-to-shoot translocation capacities than seminal roots, whereas no differences were found at micromolar nitrate supply. Since these marked differences were not reflected by the transcript levels of low-affinity nitrate transporter genes, we hypothesize that the larger metaxylem volume of nodal roots enhances predominantly the low-affinity uptake and translocation capacities of nutrients that are transported with the bulk flow of water, like nitrate

Nitrogen isotope signature evidences ammonium deprotonation as a common transport mechanism for the AMT-Mep-Rh protein superfamily

Ammonium is an important nitrogen (N) source for living organisms, a key metabolite for pH control, and a potent cytotoxic compound. Ammonium is transported by the widespread AMT-Mep-Rh membrane proteins, and despite their significance in physiological processes, the nature of substrate translocation (NH3/NH4+) by the distinct members of this family is still a matter of controversy. Using Saccharomyces cerevisiae cells expressing representative AMT-Mep-Rh ammonium carriers and taking advantage of the natural chemical-physical property of the N isotopic signature linked to NH4+/NH3 conversion, this study shows that only cells expressing AMT-Mep-Rh proteins were depleted in N-15 relative to N-14 when compared to the external ammonium source. We observed N-15 depletion over a wide range of external pH, indicating its independence of NH3 formation in solution. On the basis of inhibitor studies, ammonium transport by nonspecific cation channels did not show isotope fractionation but competition with K+. We propose that kinetic N isotope fractionation is a common feature of AMT-Mep-Rh-type proteins, which favor N-14 over N-15, owing to the dissociation of NH4+ into NH3+ H+ in the protein, leading to N-15 depletion in the cell and allowing NH3 passage or NH3/H+ cotransport. This deprotonation mechanism explains these proteins' essential functions in environments under a low NH4+/K+ ratio, allowing organisms to specifically scavenge NH4+. We show that N-15 isotope fractionation may be used in vivo not only to determine the molecular species being transported by ammonium transport proteins, but also to track ammonium toxicity and associated amino acids excretion.I. A. was supported by a postdoctoral fellowship from the Government of Navarra, Spain (Anabasid outgoing Programme, 2011) and by a postdoctoral fellowship from the Portuguese Fundaçao para a Ciencia e a Tecnologia (SFRH/BPD/90436/2012). A.M.M. is a senior research associate of the Belgian Fonds de la Recherche Scientifique Fonds de la Recherche Scientifique-FNRS (grants CDR J017617F, PDR T011515F, and ARC) and a WELBIO investigator, and M.B. is a scientific research worker supported by WELBIO. This work was also developed in the context of the following projects: PTDC/BIA-BEC/099323/2008 and PTDC/AGR-PRO/115888/2009 to cE3c and FCUL, UID/DTP/04138/2013 to iMed. ULisboa, and AGL2015-64582-C3-1-R and AGL2012-37815-C05-05 to UPNa

The Vacuolar Manganese Transporter MTP8 Determines Tolerance to Iron Deficiency-Induced Chlorosis in Arabidopsis

Iron (Fe) deficiency is a widespread nutritional disorder on calcareous soils. To identify genes involved in the Fe deficiency response, Arabidopsis (Arabidopsis thaliana) transfer DNA insertion lines were screened on a high-pH medium with low Fe availability. This approach identified METAL TOLERANCE PROTEIN8 (MTP8), a member of the Cation Diffusion Facilitator family, as a critical determinant for the tolerance to Fe deficiency-induced chlorosis, also on soil substrate. Subcellular localization to the tonoplast, complementation of a manganese (Mn)-sensitive Saccharomyces cerevisiae yeast strain, and Mn sensitivity of mtp8 knockout mutants characterized the protein as a vacuolar Mn transporter suitable to prevent plant cells from Mn toxicity. MTP8 expression was strongly induced on low-Fe as well as high-Mn medium, which were both strictly dependent on the transcription factor FIT, indicating that high-Mn stress induces Fe deficiency. mtp8 mutants were only hypersensitive to Fe deficiency when Mn was present in the medium, which further suggested an Mn-specific role of MTP8 during Fe limitation. Under those conditions, mtp8 mutants not only translocated more Mn to the shoot than did wild-type plants but suffered in particular from critically low Fe concentrations and, hence, Fe chlorosis, although the transcriptional Fe deficiency response was up-regulated more strongly in mtp8. The diminished uptake of Fe from Mn-containing low-Fe medium by mtp8 mutants was caused by an impaired ability to boost the ferric chelate reductase activity, which is an essential process in Fe acquisition. These findings provide a mechanistic explanation for the long-known interference of Mn in Fe nutrition and define the molecular processes by which plants alleviate this antagonism

Metal Tolerance Protein 8 Mediates Manganese Homeostasis and Iron Reallocation during Seed Development and Germination

Metal accumulation in seeds is a prerequisite for germination and establishment of plants but also for micronutrient delivery to humans. To investigate metal transport processes and their interactions in seeds, we focused on METAL TOLERANCE PROTEIN8 (MTP8), a tonoplast transporter of the manganese (Mn) subclade of cation diffusion facilitators, which in Arabidopsis (Arabidopsis thaliana) is expressed in embryos of seeds. The x-ray fluorescence imaging showed that expression of MTP8 was responsible for Mn localization in subepidermal cells on the abaxial side of the cotyledons and in cortical cells of the hypocotyl. Accordingly, under low Mn availability, MTP8 increased seed stores of Mn, required for efficient seed germination. In mutant embryos lacking expression of VACUOLAR IRON TRANSPORTER1 (VIT1), MTP8 built up iron (Fe) hotspots in MTP8-expressing cells types, suggesting that MTP8 transports Fe in addition to Mn. In mtp8 vit1 double mutant seeds, Mn and Fe were distributed in all cell types of the embryo. An Fe transport function of MTP8 was confirmed by its ability to complement Fe hypersensitivity of a yeast mutant defective in vacuolar Fe transport. Imbibing mtp8-1 mutant seeds in the presence of Mn or subjecting seeds to wet-dry cycles showed that MTP8 conferred Mn tolerance. During germination, MTP8 promoted reallocation of Fe from the vasculature. These results indicate that cell type-specific accumulation of Mn and Fe in seeds depends on MTP8 and that this transporter plays an important role in the generation of seed metal stores as well as for metal homeostasis and germination efficiency under challenging environmental conditions

Canonical Analysis (CA) of bacterial transcript levels and vector fitting of the root exudate metabolites.

<p>Blue arrows represent significant fittings (P<0.05). Green ellipses depict biological replicates of nutritional deficiency treatments applied to maize plants from which root exudates were collected. N = Nitrogen deficiency, P = Phosphate deficiency, Fe = Iron deficiency, and K = Potassium deficiency. Three small circles located inside each ellipse represent biological replicates for each treatment. Red crosses represent gene coordinates.</p

Heat map depicting relative transcript levels of differentially expressed genes in FZB42 exposed to root exudates collected from maize cultivated under nitrogen (N), phosphate (P), iron (Fe), and potassium (K) deficiencies in the transient growth phase (OD 3.0).

<p>Only genes that have been referred in in the ‘<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0068555#s3" target="_blank">Results and Discussion</a>’ section are displayed, the full list of differentially expressed genes is shown in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0068555#pone.0068555.s002" target="_blank">Tables S2</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0068555#pone.0068555.s004" target="_blank">S4</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0068555#pone.0068555.s006" target="_blank">S6</a>, and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0068555#pone.0068555.s008" target="_blank">S8</a>.</p

Number of differentially expressed genes of FZB42 in exponential (OD 1.0) and transient (OD 3.0) growth phases in response to nutrient-deficient maize root exudates treatments.

<p>‘-N’ denotes nitrogen deficiency; ‘-P’, phosphorus deficiency, ‘-Fe’, iron deficiency; ‘-K’, potassium deficiency.</p

Heat map showing transcript levels of the most discriminating bacterial genes for nitrogen (N), phosphorus (P), iron (Fe), and potassium (K) deficiency exudate treatments.

<p>Designations of treatments on the left of the heat maps (N, P, Fe, or K) correspond to treatments which the described gene is discriminating for. The designations for non-coding RNAs ncRNA_3, ncRNA_6, and ncRNA_2 correspond to gene IDs FZB42_3984, FZB42_4030, and FZB42_3912, respectively.</p