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First published online October 31, 2008; 10.1104/pp.108.129866 Plant Physiology 149:549-560 (2009) © 2009 American Society of Plant Biologists Sulfur Transfer through an Arbuscular Mycorrhiza1Department of Plant Biology, Michigan State University, East Lansing, Michigan 48824
Despite the importance of sulfur (S) for plant nutrition, the role of the arbuscular mycorrhizal (AM) symbiosis in S uptake has received little attention. To address this issue, 35S-labeling experiments were performed on mycorrhizas of transformed carrot (Daucus carota) roots and Glomus intraradices grown monoxenically on bicompartmental petri dishes. The uptake and transfer of 35SO42– by the fungus and resulting 35S partitioning into different metabolic pools in the host roots was analyzed when altering the sulfate concentration available to roots and supplying the fungal compartment with cysteine (Cys), methionine (Met), or glutathione. Additionally, the uptake, transfer, and partitioning of 35S from the reduced S sources [35S]Cys and [35S]Met was determined. Sulfate was taken up by the fungus and transferred to mycorrhizal roots, increasing root S contents by 25% in a moderate (not growth-limiting) concentration of sulfate. High sulfate levels in the mycorrhizal root compartment halved the uptake of 35SO42– from the fungal compartment. The addition of 1 mM Met, Cys, or glutathione to the fungal compartment reduced the transfer of sulfate by 26%, 45%, and 80%, respectively, over 1 month. Similar quantities of 35S were transferred to mycorrhizal roots whether 35SO42–, [35S]Cys, or [35S]Met was supplied in the fungal compartment. Fungal transcripts for putative S assimilatory genes were identified, indicating the presence of the trans-sulfuration pathway. The suppression of fungal sulfate transfer in the presence of Cys coincided with a reduction in putative sulfate permease and not sulfate adenylyltransferase transcripts, suggesting a role for fungal transcriptional regulation in S transfer to the host. A testable model is proposed describing root S acquisition through the AM symbiosis.
Plants and arbuscular mycorrhizal (AM) fungi have been coevolving since the Devonian period (Simon et al., 1993  ; Brundrett, 2002). Today, the AM symbiosis involves an estimated 80% of land plant species and 92% of plant families (Wang and Qiu, 2006). This symbiosis augments nutrient uptake by roots, which deplete the rhizosphere at a rate dependent on water availability, as well as the concentration and soil mobility of the nutrient (Bhat et al., 1976; Li et al., 1991; Gahoonia et al., 1994; Wang et al., 2004). Phosphorus (P) is particularly immobile in soils (Gahoonia and Nielsen, 1991), and the AM symbiosis can increase plant growth in P-deficient soils 19-fold (Hayman and Mosse, 1971). Although P acquisition is a major benefit of the symbiosis, a growing body of research points toward a more complex role for arbuscular mycorrhizas in nutrient uptake (Liu et al., 2002; Lambers et al., 2008). Nitrogen (N) nutrition, for instance, is improved in a number of crop species (Ortiz-Ceballos et al., 2007; Perner et al., 2008), and a monoxenic culture system consisting of transformed roots in symbiosis with the mycorrhizal fungus Glomus intraradices has been used to demonstrate the capacity of this symbiosis to transfer N (Govindarajulu et al., 2005; Jin et al., 2005).
Plants take up sulfur (S) primarily as the sulfate anion (Leustek, 1996
The small number of reports on the effects of AM colonization on the uptake of S, which is present in amount equal to P in plants, have presented differing results (Gray and Gerdemann, 1973
Though little is known about the mechanism of S assimilation and its regulation in mycorrhizal fungi, these pathways have been extensively characterized in other fungal species (for review, see Marzluf, 1997 This study was aimed at determining the forms of S taken up, metabolized, and transferred to its host roots by the fungal partner in the AM symbiosis. A second goal was to identify effectors of S transfer to host roots and to begin to identify AM fungal genes of S handling and their regulation. Using an AM symbiosis of monoxenic cultured roots, the transfer of S was shown to be nutritionally significant and to be regulated by S availability to the host and by S metabolite availability to the extraradical mycelia (ERM). The uptake and transfer of reduced S through the symbiosis was also demonstrated. Putative homologs of S uptake and metabolism genes were also identified and their transcriptional regulation examined, indicating that S uptake is transcriptionally regulated at the level of sulfate permease. These findings allow the development of a testable model of S uptake and transfer in the AM symbiosis.
Growth and Sulfate Uptake by Nonmycorrhizal Roots To establish conditions for studying S transfer between host roots and AM fungus, the relationship between sulfate availability, root growth, and sulfate utilization was investigated in uncolonized roots (Fig. 1 ). In the absence of available sulfate, there was a 3-fold decrease in root growth compared to growth under saturating sulfate levels (3 mM; Fig. 1A). Final dry weights were not significantly different when compared singly at sulfate concentrations above 0.005 mM; however, there was a consistent trend of increasing dry weight with increased S availability. Root growth was detected even in roots exposed to medium containing no S, probably due to internal S stores within the initial root segments. A much greater difference was seen in the uptake and incorporation of sulfate when roots were labeled with 35SO4–2 at high (3 mM), low (0.014 mM), or intermediate (0.114 mM) sulfate levels (Fig. 1B). There was a close to 4-fold increase in both aqueous alcohol-soluble and -insoluble S content from low to intermediate S conditions and an 11-fold increase between low and high S. A reduction in the root compartment sulfate concentration from 3 mM to 0.12 mM resulted in a reduction in uptake of 64% (Fig. 1B), with little or no reduction (27% difference in mean values, not statistically significant at the 95% confidence level) in growth (Fig. 1A). Lowering the sulfate available to mycorrhizal roots to <0.05 mM had a negative impact on the symbiosis, reducing the number of plates where the fungal ERM grew over the barrier from >80% to <20% (data not shown). There was no such reduction when 0.12 mM sulfate was utilized.
Fungal Uptake and Transfer of Sulfate Is Influenced by Root Access to Sulfate The ERM of G. intraradices takes up sulfate at significant rates (Fig. 2A ) and transfers most of it to the mycorrhizal roots (Fig. 2B). The total amount of 35S in ERM tissue was <5% of the amount found in the root tissue because the root biomass is >20 times the fungal biomass (tissue mass data not shown). The availability of S in the root compartment strongly influences fungal uptake in the distal ERM compartment. More than twice as much sulfate was removed from the fungal compartment when the corresponding root compartment was exposed to 0.12 mM versus 3 mM S (Fig. 2A). When roots were grown in 0.12 mM sulfate, uptake by the fungus was linear up to day 40 (R2 = 0.98). The fungus removed 0.98 µg of S from the media per day during this time. Fungal uptake of S when 3 mM sulfate was supplied to roots was linear (0.83 µg/d of S; R2 = 0.92) only for the first 20 d before the rate of removal slowed.
The total amount of S transferred from the fungal compartment to roots grown at high sulfate levels was about one-quarter of that in roots exposed to moderate sulfate levels (Fig. 2B). In contrast, there was no significant effect of sulfate levels in the root compartment on the incorporation of S by the distal ERM. Additionally, in roots growing at moderate S levels, the fraction containing sulfate and the S amino acids contained approximately twice as much 35S as did the same fraction in the ERM. The amount of 35S in the media of the colonized root compartments was monitored throughout the experiment (see "Materials and Methods") and was found to be low. The movement into the root compartment media from the fungal compartment was linear (R2 = 0.98) at 63 ng/d S when the root compartment media contained 0.12 mM sulfate. This represents 6.4% of the amount transferred to the roots. The movement into the high S root media was also linear (R2 = 0.99), with 55 ng/d S being moved across the barrier, representing 6.6% of the amount transferred to the roots.
The relative contribution of S obtained from the fungal ERM to total root S was determined at intermediate S levels (0.22 mM) by comparing fungal-derived 35S in roots with 35S directly absorbed by the roots themselves. Plates were labeled with 35SO42– in the root compartment to determine the amount of S incorporated by direct root uptake. Half of these plates were also labeled in the fungal compartment with 35SO42– at 0.22 mM sulfate, while no S was added to the fungal compartments of the remaining plates. The roots took up most of the labeled sulfate initially provided within 8 weeks (Fig. 3A ). This uptake was not affected by the presence of sulfate in the fungal compartments. The uptake of S from the fungal compartment media began when the fungus crossed the barrier (approximately 30 d after inoculation) and was comparable to direct root uptake rates in the next few weeks (Fig. 3A). S incorporation by the root and fungal tissue after approximately 10 weeks of growth is shown in Figure 3, B and C. When the fungal compartment contained 35SO42–, the total incorporation of S by the roots increased by approximately 25% (Fig. 3B). The largest change was in the sulfate fraction, which increased by 60%. There were no significant differences in root growth observed when sulfate was added to the fungal compartments (root weight data not shown), which is consistent with the lack of significant growth increase between moderate and high sulfate growth conditions (Fig. 1). When the fungal side was labeled, total incorporation of 35S by the fungus was similar in ERM collected from the fungal and root compartments when the fungal side was labeled (Fig. 3C), which is consistent with the absence of any known physiological differences between the ERM in the two compartments, and showing that the availability of S to the distal ERM did not affect incorporation by the ERM in the root compartment. 35S accumulated in the fungal ERM from the unlabeled fungal compartment, showing that in a low S environment (0.02 mM), S can be moved by the fungus from the root compartment to the distal ERM (Fig. 3C).
The Effect of S Metabolites on S Uptake and Transfer through the Mycorrhizal Symbiosis
Radiolabeled sulfate was added to the fungal compartments of split plates together with Met, Cys, and GSH in amounts previously shown to suppress sulfate assimilation in other fungi (Ketter and Marzluf, 1988
GSH greatly reduced the growth of fungal ERM in the distal compartment, though not the root growth (not shown), so it is unclear whether the reduction in the transfer of S by GSH-treated ERM is due to the regulation of sulfate uptake or simply to reduced fungal growth. Neither Cys nor Met significantly affected the growth of fungal ERM or roots, with ERM biomass approximately doubling between weeks 2 and 4 (data not shown). Thus, the effect of Cys on S transfer to mycorrhizal roots does not appear to be due to growth inhibition.
Fungal ERM imports and transfers substantial amounts of reduced S supplied as Cys or Met (Fig. 5 ). The amount of S transferred when 35S-labeled Cys is the sole source of S in the fungal compartment is comparable to the S transfer when plates are labeled with sulfate (Figs. 2B, 3B, and 4, A–C). Roughly one-half of this amount was transferred when Met was provided (Fig. 5). The incorporation of S by the fungal mycelium per milligram dry weight when labeled Cys is supplied to the fungal compartment is about 40% less than the level found in the roots. Utilizing Met as an S source led to a 46% increase in the amino acid fraction in the ERM (Fig. 5). There were striking differences between the relative amount of S in the solubilized fractions in both roots and fungal mycelia when reduced S was supplied compared to sulfate. Labeling with Cys and Met resulted in a much higher percentage of labeling in the solubilized root tissue pool (78% and 88%, respectively) than when sulfate was supplied as the primary S source (7%–21%; Fig. 4, A–D). The opposite relationship was found in fungal mycelia, where applying Cys or Met as primary sources of S led to solubilized tissue fractions that were at most 8% of the total S incorporated (Fig. 5). When labeled sulfate was the primary S source, 5% to 22% of the total S incorporation is found in the tissue solubilized fraction (Fig. 4, D–F).
Gene Expression Measurements
The expression of fungal transcripts encoding putative genes involved in sulfate assimilation was analyzed in relation to the application of Cys to the distal ERM. Partial sequences of transcripts representing putative S assimilatory genes were identified by high throughput 454 sequencing of cDNA from fungal ERM grown in split plates in M medium (3 mM sulfate). These included a 234-bp sequence with 67% identity at the amino acid level to SUL1 from S. cerevisiae and a 247-bp sequence with 67% and 70% identities at the amino acid level to S adenylyltransferase from A. nidulans and A. terreus, respectively. Sequences with homology to enzymes defining the reverse trans-sulfuration pathway were also identified. These included a 403-bp sequence with 79% and 71% identities at the amino acid level to cystathionine β-synthase from A. fumigatus and CYS4 from S. cerevisiae. Additionally, a 220-bp sequence was identified with 58% and 69% amino acid identities to CYS3 from S. cerevisiae and cystathionine
The uptake of 35S by nonmycorrhizal roots was dependent on the external sulfate concentration (Fig. 1B), as has been reported for tomato (Solanum lycopersicum) seedlings and carrot (Daucus carota) storage root sections (Cram, 1983 ; Lopez et al., 2002). In carrot, Cram et al. (1983) showed a proportional increase up to an external concentration of 50 mM, leading the authors to conclude that feedback inhibition of sulfate uptake is absent. Vegetative root growth was less affected by alterations in available sulfate concentrations than was uptake or reduction (Fig. 1A). Subsequent measurements of mycorrhizal uptake activity were conducted using sulfate concentrations that increased potential differences in root uptake and contents and minimized the limitation of growth by S.
Bicompartmental petri plate cultures are a well-established model mycorrhizal system (St. Arnaud et al., 1996) used extensively in studies of mycorrhizal metabolism and nutrient transfer (e.g. Govindarajulu et al., 2005 After supplying 35S-sulfate to the fungal compartment under a range of conditions, labeled sulfate was invariably found in roots in amounts that dwarfed the total quantities in the fungal mycelium. For example, after 6 weeks of labeling the fungal compartment with 35SO42–, total sulfate in the fungus was 5.1% of the amount in the roots (Fig. 4, C and F). Roots in that experiment were grown in 0.5 mM sulfate, and the distal ERM was supplied with 0.12 mM, and the same was true (5.3%) when the roots were grown in 0.22 mM sulfate and the fungus was also labeled with 0.22 mM sulfate (Fig. 3, B and C). Based on this evidence, one may conclude that the sulfate anion is transferred by G. intraradices to host roots. However, as shown in Figure 5, the fungus is clearly also capable of the uptake and transfer of reduced forms of S at rates comparable to sulfate. Thus, the ERM can supply the host plant with organic forms of S from the soil. Because an estimated 95% of the S in soils is in organic forms, the ability of mycorrhizal plants to access S from this source is potentially important for plant nutrition.
The first steps toward understanding the regulation of sulfate uptake by a mycorrhizal fungus in the symbiotic state were taken by analyzing the effect of common S metabolite repressors Cys, Met, and GSH on the uptake and transfer of radiolabeled sulfate through the fungus. The addition of 1 mM met was shown to suppress the expression of S assimilation pathway genes in yeast (Kuras and Thomas, 1995
By contrast, the application of 1 mM Cys to the ERM resulted in S transfer from fungus to mycorrhizal roots being approximately halved (Fig. 4, A–C). The amount by which sulfate transfer is reduced in the presence of Cys (Fig. 4) is similar to the level of S that is taken up as Cys and transferred to the roots (Fig. 5). Labeled GSH has been shown to be imported by ectomycorrhizal oak (Quercus rober) trees at rates comparable to sulfate (Seegmuller and Rennenberg, 2002
The changes in expression of genes putatively involved in sulfate transport and assimilation indicates a role for transcriptional regulation of sulfate permease in the reduction in sulfate transport in the presence of Cys. The reduction in S transfer of 45% with the application of 1 mM Cys to distal ERM (Fig. 4C) correlated with a 3.8- ± 1.2-fold reduction in the expression of mRNA encoding a putative sulfate permease. The lack of a significant change in the expression of a cDNA sequence with high homology to S-adenosyl transferases suggests that the reduction in S transfer is a function of reduced uptake rather than sulfate reduction. However, the analysis of more S assimilation pathway genes and compartment-specific sulfate permease isozymes is needed before concluding that the transcriptional regulation of this sulfate permease gene explains the reduction in S transfer with the addition of Cys. Sequences encoding putative genes with homology to cystathionine β-synthase and cystathionine Figure 6 presents a working model consistent with the findings presented. We have demonstrated uptake, assimilation, and transfer of sulfate (Figs. 3B and 4, A–C) and of reduced S (Fig. 5) by the fungal partner to the host roots in physiologically significant quantities, along with bidirectional transport of sulfate within the fungal ERM (Fig. 3C). The inverse relationship between the rhizospheric sulfate concentration and the amount of sulfate uptake from the fungal compartment (Fig. 2, A and B) suggests that an increase in root intracellular or apoplastic sulfate can suppress both the transfer of sulfate from the intraradical hyphae to the host and the uptake by the extraradical hyphae. This relationship is depicted in the model as the negative regulation of sulfate transfer across the root-hyphal interface by root sulfate levels, which subsequently results in an increase in the fungal intracellular sulfate concentration and suppression of uptake in the distal ERM. The relationship between the transfer of a reduced form of S when Cys or Met are imported by the fungus and the uptake of sulfate in the presence of Cys and Met suggest a simultaneous transfer of sulfate and reduced S and/or a regulatory response. Relative real-time PCR measurements of expression for a putative high affinity sulfate permease gene revealed an approximately 73% reduction in transcript levels when the ERM was exposed to 1 mM Cys, indicating a regulatory effect on sulfate uptake (Figs. 6 and 7 ). The working model proposed is a testable interpretation of the findings presented that invites future experiments to verify gene identities, localize expression within the host roots, and analyze regulation in mycorrhizas of whole plants.
Chemicals and Reagents
Gel-Gro gellan (MP Biomedical) was used for the solidification of growth media. Radioactively labeled sulfate was obtained as Na235SO4 from MP Biomedicals. Labeled Cys and Met were separated from a crude mixture of 35S-labeled compounds (TRAN35S-LABEL; MP Biomedicals) containing
All experimental procedures used M medium (Fortin et al., 2002
Uncolonized Ri T-DNA-transformed roots of carrot (Daucus carota; DC2; Diop et al., 1992
Root material was collected with forceps and rinsed for 5 min in deionized water to remove external, and reduce apoplastic, 35S. The collected roots were rinsed again with deionized water in a separate container, blotted dry, frozen in liquid N, and lyophilized. ERM was collected by blending the solidified medium at high speed in 10 mM sodium citrate buffer, pH 6.0, at an approximate gel:buffer ratio of 1:2.5 for 2 min, which dissolved the gellan (Pfeffer et al., 1999
For biochemical fractionation, lyophilized fungal mycelium was pulverized with two 3-mm stainless steel beads using a bead mill (Retsch MM301). Due to the high lipid content, 0.1 mL of cold methanol:water (70:30) was added to aid in disruption. The samples were shaken at 30 Hz for 4 min, and 2-µL samples were analyzed by dissecting microscope to ensure that hyphae and any spores had been broken. After disruption, 0.9 mL of cold methanol:water (70:30) was added, and the sample was vortexed for 5 min. Samples were then centrifuged and supernatants collected. The cold aqueous methanol extraction was repeated twice more using 1 mL each time and the supernatants pooled. Then 0.5 mL of supernatant solution was scintillation counted after adding to 5 mL of BioSafe II (MP Biomedicals) scintillation cocktail. To determine the amount of sulfate in the samples, the sulfate was precipitated from an aliquot of the aqueous alcohol extract by adding 0.1 mL of 100 mM Na2SO4 and 0.3 mL of 10 mM HCl solution to 0.5 mL of the sample followed by vortexing and the addition of 0.1 mL of 100 mM BaCl2. Samples were then incubated for 30 min at 100°C. The resulting barium sulfate precipitate was removed by centrifugation, and the sulfate-free supernatant was scintillation counted. The amount of sulfate in each sample was determined by the subtraction of these counts from those obtained from an aliquot of the original aqueous alcohol extract. Tests with standards showed that 99.8% ± 0.1% of sulfate was removed by this procedure (three trials with three samples each) and that 96.4% ± 1.3% of 35S-Cys and 95.6% ± 1.6% of 35S-Met remained in solution.
Residues after aqueous ethanol extraction were extracted using 1 mL of protein extraction buffer containing 9 M urea, 1% SDS, 25 mM Tris-HCl, pH 6.8, 1 mM EDTA, and 0.7 M 2-mercaptoethanol as described by Osherov and May (1998) Dried roots were pulverized in 15-mL centrifuge tubes with 10 5-mm metal beads for 30 min using a paint shaker. Five milliliters of cold methanol:water (70:30) was added to the powder, and the tubes were vortexed for 5 min. While keeping particulates suspended, a 1-mL aliquot of the solution was transferred to a microcentrifuge tube and centrifuged. Subsequent steps were as for fungal samples.
Three 1-cm diameter wells were excavated in the gel at the time of labeling with a sterilized cork borer as far from one another as possible. Then 0.1 to 0.2 mL of media of the same composition as that in the compartment being analyzed was added to each well, and this volume was maintained through weekly refilling as needed. At the start of the experiment, label was added to one of the wells, and 35S in each of the three wells were measured after 1 week to ensure efficient diffusion of the radioactivity through the plate. 35S contents were measured by adding 50 µL of liquid media to 5 mL of scintillation fluid, followed by scintillation counting.
Roots were grown on solidified M media containing 0.12 mM S as Na2SO4 for 4 weeks, and approximately 5-cm root segments were aseptically removed, weighed, and added to 20 mL of liquid M media without S (ZnSO4 7H2O and CuSO4 5H2O replaced by 1.2 mg of ZnCl and 0.5 mg of CuCl2) for measurements of S-mediated growth limitation. For each sulfate concentration (0, 0.005, 0.01, 0.02, 0.05, 0.1, 0.5, and 3 mM), roots were weighed and then placed on 20 mL of solidified media per side of three bicompartmental petri plates and allowed to grow for 3 months. The roots were collected, dried at 70°C overnight, and weighed. To measure S uptake, roots were grown at 0.12 mM sulfate as above and transferred into 20 mL of S media solidified with gellan containing 0.02, 0.12, or 3 mM sulfate labeled with 11.5 µCi of Na2[35SO4]. After 4 weeks, the roots were removed from the media, rinsed several times in deionized water to remove external radioactivity, lyophilized, and weighed. The extraction procedure for the radiolabeled tissue was simplified to include only the aqueous alcohol and solubilization steps.
Cys, Met, and GSH were added to a final concentration of 1 mM (as 0.2–0.5 mL of sterile aqueous solutions) to the fungal compartments of split plates with 10-week-old roots growing with 0.52 mM Na2SO4. The uptake of 37.4 µCi of Na2[35SO4], which was added to fungal compartments simultaneously with metabolic regulators and 0.12 mM Na2SO4, was monitored by analyzing aliquots of media from a liquid-filled well as described above. The plates were collected at 2, 4, and 6 weeks. The fungal mycelium was collected by dissolving the gellan in a blender with 10 mM sodium citrate, pH 6, then sieving the blended solution. The roots were extracted from the media using forceps and rinsed in deionized water for 5 min before freezing. The fungal and root tissues were extracted as described above. Using cultures grown as for the above experiment, 1 mM of Cys labeled with 25.7 µCi of [35S]Cys or 1 mM Met labeled with 54.2 µCi of [35S]Met was applied to the fungal sides of 15 plates as the sole S source. The plates were incubated at 25°C for 1 month prior to collecting and extracting the biochemical fractions as described above.
Sequences of putative S gene fragments were identified from an EST database (Jun et al., 2002
Mycorrhizal split plates were grown until the fungal compartment was approximately one-half colonized. To the fungal compartment, 1 mL of a filter sterilized solution containing either sodium sulfate or sodium sulfate and Cys was applied to give a final concentration of 0.12 mM sulfate and 1 mM Cys. Plates were incubated for 24 h before tissue from five to seven plates was collected as described above and immediately frozen in liquid N. RNA was extracted and converted to cDNA as described above. The initial quantitative real-time PCR (qRT-PCR) reaction mixture containing primers at a concentration of 300 nM and 1 ng of cDNA template was made to 0.015 mL in a 96-well plate, 0.01 mL of which was transferred to a 384-well plate, and the PCR reactions were monitored using an ABI Prism 7900 HT Sequence Detection system (Applied Biosystems) with the following cycling program: 50°C for 2 min, 95°C for 10 min, followed by 40 cycles of 95°C for 15 s, and 60°C for 1 min. Power SYBR Green 2-Step Master mix (Applied Biosystems) was used for all real-time PCR assays. The
The sequences studied were submitted to GenBank and given the following accession numbers: putative high affinity sulfate permease (FJ161947); putative sulfate adenylyltransferase (FJ161948); putative cystathionine
We thank Inga Krassovskaya for help with molecular data analysis and protocols, Joseph Leykam for the HPLC separation of [35S]Cys and [35S]Met, and the Research Technology Support Facility at Michigan State University for sequencing and qRT-PCR analyses. Received September 12, 2008; accepted October 29, 2008; published October 31, 2008.
1 This work was supported by the National Science Foundation (award no. 0616016).  The author responsible for distribution of materials integral to the findings presented in this article in accordance with the policy described in the Instructions for Authors (www.plantphysiol.org) is: James W. Allen (allenj28{at}msu.edu). www.plantphysiol.org/cgi/doi/10.1104/pp.108.129866 * Corresponding author; e-mail allenj28{at}msu.edu.
Banerjee R, Evande R, Kabil O, Ojha S, Taoka S (2003) Reaction mechanism and regulation of cystathionine beta-synthase. Biochim Biophys Acta 1647: 30–35[Medline] Baumgardner RE, Lavery TF, Rogers CM, Isil SS (2002) Estimates of the atmospheric deposition of sulfur and nitrogen species: Clean Air Status and Trends Network, 1990-2000. Environ Sci Technol 36: 2614–2629[Medline] Bhat KKS, Nye PH, Baldwin JP (1976) Diffusion of phosphate to plant roots in soil. 4. Concentration distance profile in rhizosphere of roots with root hairs in a low-p soil. Plant Soil 44: 63–72[CrossRef][Web of Science] Bradfield G, Somerfield P, Meyn T, Holby M, Babcock D, Bradley D, Segel IH (1970) Regulation of sulfate transport in filamentous fungi. Plant Physiol 46: 720–727 Breton A, Surdin-Kerjan Y (1977) Sulfate uptake in Saccharomyces cerevisiae: biochemical and genetic study. J Bacteriol 132: 224–232 Brundrett MC (2002) Coevolution of roots and mycorrhizas of land plants. New Phytol 154: 275–304[CrossRef][Web of Science] Brzywczy J, Sienko M, Kucharska A, Paszewski A (2002) Sulphur amino acid synthesis in Schizosaccharomyces pombe represents a specific variant of sulphur metabolism in fungi. Yeast 19: 29–35[CrossRef][Web of Science][Medline] Cherest H, Davidian JC, Thomas D, Benes V, Ansorge W, Surdin-Kerjan Y (1997) Molecular characterization of two high affinity sulfate transporters in Saccharomyces cerevisiae. Genetics 145: 627–635[Abstract] Clarke DL, Newbert RW, Turner G (1997) Cloning and characterisation of the adenosyl phosphosulphate kinase gene from Aspergillus nidulans. Curr Genet 32: 408–412[CrossRef][Web of Science][Medline] Cooper KM, Tinker PB (1978) Translocation and transfer of nutrients in vesicular-arbuscular mycorrhizas. 2. Uptake and translocation of phosphorus, zinc and sulfur. New Phytol 81: 43–52[CrossRef][Web of Science] Cram J (1983) Characteristics of sulfate transport across plasmalemma and tonoplast of carrot root-cells. Plant Physiol 72: 204–211 Diop T, Becard G, Pichey Y (1992) Long-term invitro culture of an endomycorrhizal fungus, Gigaspora margarita, on Ri T-dna transformed roots of carrot. Symbiosis 12: 249–259[Web of Science] Eriksen J, Askegaard M (2000) Sulphate leaching in an organic crop rotation on sandy soil in Denmark. Agric Ecosyst Environ 78: 107–114[CrossRef] Fitzgerald JW (1976) Sulfate ester formation and hydrolysis: potentially important yet often ignored aspect of sulfur cycle of aerobic soils. Bacteriol Rev 40: 698–721 Fortin JA, Becard G, Declerck S, Dalpe Y, St-Arnaud M, Coughlan AP, Piche Y (2002) Arbuscular mycorrhiza on root-organ cultures. Can J Bot 80: 1–20[CrossRef] Gahoonia TS, Nielsen NE (1991) A method to study rhizosphere processes in thin soil layers of different proximity to roots. Plant Soil 135: 143–146[CrossRef][Web of Science] Gahoonia TS, Raza S, Nielsen NE (1994) Phosphorus depletion in the rhizosphere as influenced by soil-moisture. Plant Soil 159: 213–218[CrossRef][Web of Science] Govindarajulu M, Pfeffer PE, Jin HR, Abubaker J, Douds DD, Allen JW, Bucking H, Lammers PJ, Shachar-Hill Y (2005) Nitrogen transfer in the arbuscular mycorrhizal symbiosis. Nature 435: 819–823[CrossRef][Medline] Gray LE, Gerdemann JW (1973) Uptake of sulphur-35 by vesicular-arbuscular mycorrhizae. Plant Soil 39: 687–689[Web of Science] Grynberg M, Piotrowska M, Pizzinini E, Turner G, Paszewski A (2001) The Aspergillus nidulans metE gene is regulated by a second system independent from sulphur metabolite repression. Biochim Biophys Acta 1519: 78–84[Medline] Guo T, Zhang JL, Christie P, Li XL (2007) Pungency of spring onion as affected by inoculation with arbuscular mycorrhizal fungi and sulfur supply. J Plant Nutr 30: 1023–1034[CrossRef][Web of Science] Hansen J, Johannesen PF (2000) Cysteine is essential for transcriptional regulation of the sulfur assimilation genes in Saccharomyces cerevisiae. Mol Gen Genet 263: 535–542[CrossRef][Web of Science][Medline] Hawkins HJ, Johansen A, George E (2000) Uptake and transport of organic and inorganic nitrogen by arbuscular mycorrhizal fungi. Plant Soil 226: 275–285[CrossRef][Web of Science] Hayman DS, Mosse B (1971) Plant growth responses to vesicular-arbuscular mycorrhiza. 1. Growth of endogone-inoculated plants in phosphate-deficient soils. New Phytol 70: 19–27[CrossRef][Web of Science] Jin H, Pfeffer PE, Douds DD, Piotrowski E, Lammers PJ, Shachar-Hill Y (2005) The uptake, metabolism, transport and transfer of nitrogen in an arbuscular mycorrhizal symbiosis. New Phytol 168: 687–696[CrossRef][Web of Science][Medline] Joner EJ, Jakobsen I (1994) Contribution by 2 arbuscular mycorrhizal fungi to P-uptake by cucumber (Cucumis sativus l) from P-32 labeled organic-matter during mineralization in soil. Plant Soil 163: 203–209[CrossRef][Web of Science] Jun J, Rehrer C, Pfeffer PE, Shachar-Hill Y, Lammers PJ (2002) Expression in an arbuscular mycorrhizal fungus of genes putatively involved in metabolism, transport, the cytoskeleton and the cell cycle. Plant Soil 244: 141–148[CrossRef][Web of Science] Ketter JS, Jarai G, Fu YH, Marzluf GA (1991) Nucleotide-sequence, messenger-rna stability, and dna recognition elements of cys-14, the structural gene for sulfate permease-II in Neurospora crassa. Biochemistry 30: 1780–1787[CrossRef][Web of Science][Medline] Ketter JS, Marzluf GA (1988) Molecular-cloning and analysis of the regulation of cys-14+, a structural gene of the sulfur regulatory circuit of Neurospora crassa. Mol Cell Biol 8: 1504–1508 Kreuzwieser J, Rennenberg H (1998) Sulphate uptake and xylem loading of mycorrhizal beech roots. New Phytol 140: 319–329[CrossRef][Web of Science] Kuras L, Thomas D (1995) Functional-analysis of met4, a yeast transcriptional activator responsive to S-adenosylmethionine. Mol Cell Biol 15: 208–216 Lambers H, Raven JA, Shaver GR, Smith SE (2008) Plant nutrient-acquisition strategies change with soil age. Trends Ecol Evol 23: 95–103[CrossRef][Medline] Lefohn AS, Husar JD, Husar RB (1999) Estimating historical anthropogenic global sulfur emission patterns for the period 1850-1990. Atmos Environ 33: 3435–3444 Leustek T (1996) Molecular genetics of sulfate assimilation in plants. Physiol Plant 97: 411–419[CrossRef] Leustek T, Martin MN, Bick JA, Bick JA, Davies JP (2000) Pathways and regulation of sulfur metabolism revealed through molecular and genetic studies. Annu Rev Plant Physiol 51: 145–165 Li XL, George E, Marschner H (1991) Extension of the phosphorus depletion zone in VA-mycorrhizal white clover in a calcareous soil. Plant Soil 136: 41–48[CrossRef][Web of Science] Liu A, Hamel C, Elmi A, Costa C, Ma B, Smith DL (2002) Concentrations of K, Ca and Mg in maize colonized by arbuscular mycorrhizal fungi under field conditions. Can J Plant Sci 82: 271–278 Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)). Methods 25: 402–408[CrossRef][Web of Science][Medline] Lopez J, Bell CI, Tremblay N, Dorais M, Gosselin A (2002) Uptake and translocation of sulphate in tomato seedlings in relation to sulphate supply. J Plant Nutr 25: 1471–1485[CrossRef][Web of Science] Mansouri-Bauly H, Kruse J, Sykorova Z, Scheerer U, Kopriva S (2006) Sulfur uptake in the ectomycorrhizal fungus Laccaria bicolor S238N. Mycorrhiza 16: 421–427[CrossRef][Web of Science][Medline] Marzluf GA (1993) Regulation of sulfur and nitrogen-metabolism in filamentous fungi. Annu Rev Microbiol 47: 31–55[CrossRef][Web of Science][Medline] Marzluf GA (1997) Molecular genetics of sulfur assimilation in filamentous fungi and yeast. Annu Rev Microbiol 51: 73–96[CrossRef][Web of Science][Medline] Natorff R, Sienko M, Brzywczy J, Paszewski A (2003) The Aspergillus nidulans metR gene encodes a bZIP protein which activates transcription of sulphur metabolism genes. Mol Microbiol 49: 1081–1094[CrossRef][Web of Science][Medline] Ono B, Hazu T, Yoshida S, Kawato T, Shinoda S, Brzvwczy J, Paszewski A (1999) Cysteine biosynthesis in Saccharomyces cerevisiae: a new outlook on pathway and regulation. Yeast 15: 1365–1375[CrossRef][Web of Science][Medline] Ono B, Heike C, Yano Y, Inoue T, Naito K, Nakagami S, Yamane A (1992a) Cloning and mapping of CYS4 gene of Saccharomyces cerevisiae. Curr Genet 21: 285–289[CrossRef][Web of Science][Medline] Ono B, Tanaka K, Naito K, Heike C, Shinoda S, Yamamoto S, Ohmori S, Oshima T, Toh-e A (1992b) Cloning and characterization of CYS3(CYI1) gene of Saccharomyces cerevisiae. J Bacteriol 174: 3339–3347 Ono BI, Naito K, Shirahige YI, Yamamoto M (1991) Regulation of cystathionine gamma-lyase in Saccharomyces cerevisiae. Yeast 7: 843–848[CrossRef][Web of Science][Medline] Ortiz-Ceballos AI, Pena-Cabriales JJ, Fragoso C, Brown GG (2007) Mycorrhizal colonization and nitrogen uptake by maize: combined effect of tropical earthworms and velvet bean mulch. Biol Fertil Soils 44: 181–186[CrossRef] Osherov N, May GS (1998) Optimization of protein extraction from Aspergillus nidulans for gel electrophoresis. Fungal Genet Rep 45: 41–42 Paszewski A, Grabski J (1975) Enzymatic lesions in methionine mutants of Aspergillus nidulans: role and regulation of an alternative pathway od cysteine and methionine synthesis. J Bacteriol 124: 893–904 Perner H, Rohn S, Driemel G, Batt N, Schwarz D, Kroh LW, George E (2008) Effect of nitrogen species supply and mycorrhizal colonization on organosulfur and phenolic compounds in onions. J Agric Food Chem 56: 3538–3545[CrossRef][Web of Science][Medline] Pfeffer PE, Douds DD, Becard G, Shachar-Hill Y (1999) Carbon uptake and the metabolism and transport of lipids in an arbuscular mycorrhiza. Plant Physiol 120: 587–598 Pilsyk S, Natorff R, Sienko M, Paszewski A (2007) Sulfate transport in Aspergillus nidulans: a novel gene encoding alternative sulfate transporter. Fungal Genet Biol 44: 715–725[CrossRef][Web of Science][Medline] Rennenberg H (1999) The significance of ectomycorrhizal fungi for sulfur nutrition of trees. Plant Soil 215: 115–122[CrossRef][Web of Science] Rennenberg H, Herschbach C, Haberer K, Kopriva S (2007) Sulfur metabolism in plants: Are trees different? Plant Biol 9: 620–637[CrossRef][Medline] Rhodes LH, Gerdemann JW (1978a) Influence of phosphorus nutrition on sulfur uptake by vesicular-arbuscular mycorrhizae of onion. Soil Biol Biochem 10: 361–364[CrossRef] Rhodes LH, Gerdemann JW (1978b) Hyphal translocation and uptake of sulfur by vesicular-arbuscular mycorrhizae of onion. Soil Biol Biochem 10: 355–360[CrossRef] Riley NG, Zhao FJ, McGrath SP (2000) Availability of different forms of sulphur fertilisers to wheat and oilseed rape. Plant Soil 222: 139–147[CrossRef][Web of Science] Scherer HW (2001) Sulphur in crop production. Eur J Agron 14: 81–111[CrossRef] Seegmuller S, Rennenberg H (2002) Transport of organic sulfur and nitrogen in the roots of young mycorrhizal pedunculate oak trees (Quercus robur L.). Plant Soil 242: 291–297[CrossRef][Web of Science] Sienko M, Paszewski A (1999) The metG gene of Aspergillus nidulans encoding cystathionine beta-lyase: cloning and analysis. Curr Genet 35: 638–646[CrossRef][Web of Science][Medline] Simon L, Bousquet J, Levesque RC, Lalonde M (1993) Origin and diversification of endomycorrhizal fungi and coincidence with vascular land plants. Nature 363: 67–69[CrossRef][Web of Science] Spencer B, Harada T (1960) Role of choline sulphate in the sulphur metabolism of fungi. Biochem J 77: 305–315[Web of Science][Medline] St-Arnaud M, Hamel C, Vimard B, Caron M, Fortin JA (1996) Enhanced hyphal growth and spore production of the arbuscular mycorrhizal fungus Glomus intraradices in an in vitro system in the absence of host roots. Mycol Res 100: 328–332[CrossRef][Web of Science] Tabatabai MA (1986) Sulfur in Agriculture. American Society of Agronomy, Madison, WI, pp 207–226 Thomas D, Surdin-Kerjan Y (1997) Metabolism of sulfur amino acids in Saccharomyces cerevisiae. Microbiol Mol Biol Rev 61: 503–532 Wang B, Qiu YL (2006) Phylogenetic distribution and evolution of mycorrhizas in land plants. Mycorrhiza 16: 299–363[CrossRef][Web of Science][Medline] Wang ZY, Kelly JM, Kovar JL (2004) In situ dynamics of phosphorus in the rhizosphere solution of five species. J Environ Qual 33: 1387–1392 Related articles in Plant Physiol.:
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plammers/glomus/
5). DW, Dry weight.
-lyase from A. fumigatus. The presence of transcripts for these putative S genes is consistent with sulfate uptake and assimilation via the usual assimilation pathway and also strongly suggests the presence of the reverse trans-sulfuration pathway in AM fungi. The expression of the sulfate permease and reverse trans-sulfuration enzyme candidates was analyzed in ERM after exposure to 0.12 mM sulfate with or without the presence of 1 mM Cys. The conditions were chosen to enable the comparison between gene expression changes and the uptake and transfer of radiolabeled sulfate with Cys addition (
70% [35S]Met and 15% [35S]Cys. The compounds were separated using an amino acid analyzer (Hitachi L8800). Retention times for the amino acids were determined using post-column ninhydrin bonding and UV absorption. The radioactive compounds were then collected in a ninhydrin-free environment by manual collection of the appropriate fractions. 