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Symbiosis dependent accumulation of primary metabolites in arbuscule-containing cells.

Gaude N, Bortfeld S, Erban A, Kopka J, Krajinski F - BMC Plant Biol. (2015)

Bottom Line: Laser capture microdissection (LCM) combined with gas chromatography mass spectrometry (GC-EI/TOF-MS) enabled the analysis of primary metabolite levels,which might be of plant or fungal origin, within these cells.High levels of the amino acids, aspartate, asparagine, glutamate, and glutamine, were observed in arbuscule-containing cells.Elevated amounts of sucrose and the steady-state of hexose levels indicated a direct assimilation of monosaccharides by the fungal partner.

View Article: PubMed Central - PubMed

Affiliation: Max Planck Institute of Molecular Plant Physiology, Am Muehlenberg 1, 14476, Potsdam, Germany. Gaude@mpimp-golm.mpg.de.

ABSTRACT

Background: The arbuscular mycorrhizal symbiosis is characterized by the presence of different symbiotic structures and stages within a root system. Therefore tools allowing the analysis of molecular changes at a cellular level are required to reveal insight into arbuscular mycorrhizal (AM) symbiosis development and functioning.

Results: Here we describe the analysis of metabolite pools in arbuscule-containing cells, which are the site of nutrient transfer between AM fungus and host plant. Laser capture microdissection (LCM) combined with gas chromatography mass spectrometry (GC-EI/TOF-MS) enabled the analysis of primary metabolite levels,which might be of plant or fungal origin, within these cells.

Conclusions: High levels of the amino acids, aspartate, asparagine, glutamate, and glutamine, were observed in arbuscule-containing cells. Elevated amounts of sucrose and the steady-state of hexose levels indicated a direct assimilation of monosaccharides by the fungal partner.

No MeSH data available.


Workflow illustration: LCM-mediated harvest of root cortex cells for metabolite profiling. Root fragments of mycorrhizal and non-mycorrhizal Medicago truncatula plants were lyophilized and sectioned with a cryostat (a and b). In 35 μm longitudinal sections, cortical cell populations were identified and isolated by laser microdissection (c and d). Approximately 13,000 cells for each cell type (arbuscule containing cells of mycorrhizal roots [arb] and cortical cells of non-colonized roots [cor]) were collected and subjected to derivatization (e). GC-EI/TOF-MS measurements facilitated the abundance of primary metabolites in the analysed samples (f). The corresponding compounds were identified through spectral matching against the National Institute of Standards and Technology library (NIST08) (g)
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Fig1: Workflow illustration: LCM-mediated harvest of root cortex cells for metabolite profiling. Root fragments of mycorrhizal and non-mycorrhizal Medicago truncatula plants were lyophilized and sectioned with a cryostat (a and b). In 35 μm longitudinal sections, cortical cell populations were identified and isolated by laser microdissection (c and d). Approximately 13,000 cells for each cell type (arbuscule containing cells of mycorrhizal roots [arb] and cortical cells of non-colonized roots [cor]) were collected and subjected to derivatization (e). GC-EI/TOF-MS measurements facilitated the abundance of primary metabolites in the analysed samples (f). The corresponding compounds were identified through spectral matching against the National Institute of Standards and Technology library (NIST08) (g)

Mentions: For this purpose we used a modified a protocol originally developed for metabolite measurements in vascular bundle cells of A. thaliana [15] (Fig. 1). These modifications were necessary, as the highly sensitive analytical method needed a reduction of the background caused by polymeric substances originating from the fixation medium. Root fragments were longitudinally sectioned and colonized as well as non-colonized root cells were microscopically identified, cut and collected.Fig. 1


Symbiosis dependent accumulation of primary metabolites in arbuscule-containing cells.

Gaude N, Bortfeld S, Erban A, Kopka J, Krajinski F - BMC Plant Biol. (2015)

Workflow illustration: LCM-mediated harvest of root cortex cells for metabolite profiling. Root fragments of mycorrhizal and non-mycorrhizal Medicago truncatula plants were lyophilized and sectioned with a cryostat (a and b). In 35 μm longitudinal sections, cortical cell populations were identified and isolated by laser microdissection (c and d). Approximately 13,000 cells for each cell type (arbuscule containing cells of mycorrhizal roots [arb] and cortical cells of non-colonized roots [cor]) were collected and subjected to derivatization (e). GC-EI/TOF-MS measurements facilitated the abundance of primary metabolites in the analysed samples (f). The corresponding compounds were identified through spectral matching against the National Institute of Standards and Technology library (NIST08) (g)
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4590214&req=5

Fig1: Workflow illustration: LCM-mediated harvest of root cortex cells for metabolite profiling. Root fragments of mycorrhizal and non-mycorrhizal Medicago truncatula plants were lyophilized and sectioned with a cryostat (a and b). In 35 μm longitudinal sections, cortical cell populations were identified and isolated by laser microdissection (c and d). Approximately 13,000 cells for each cell type (arbuscule containing cells of mycorrhizal roots [arb] and cortical cells of non-colonized roots [cor]) were collected and subjected to derivatization (e). GC-EI/TOF-MS measurements facilitated the abundance of primary metabolites in the analysed samples (f). The corresponding compounds were identified through spectral matching against the National Institute of Standards and Technology library (NIST08) (g)
Mentions: For this purpose we used a modified a protocol originally developed for metabolite measurements in vascular bundle cells of A. thaliana [15] (Fig. 1). These modifications were necessary, as the highly sensitive analytical method needed a reduction of the background caused by polymeric substances originating from the fixation medium. Root fragments were longitudinally sectioned and colonized as well as non-colonized root cells were microscopically identified, cut and collected.Fig. 1

Bottom Line: Laser capture microdissection (LCM) combined with gas chromatography mass spectrometry (GC-EI/TOF-MS) enabled the analysis of primary metabolite levels,which might be of plant or fungal origin, within these cells.High levels of the amino acids, aspartate, asparagine, glutamate, and glutamine, were observed in arbuscule-containing cells.Elevated amounts of sucrose and the steady-state of hexose levels indicated a direct assimilation of monosaccharides by the fungal partner.

View Article: PubMed Central - PubMed

Affiliation: Max Planck Institute of Molecular Plant Physiology, Am Muehlenberg 1, 14476, Potsdam, Germany. Gaude@mpimp-golm.mpg.de.

ABSTRACT

Background: The arbuscular mycorrhizal symbiosis is characterized by the presence of different symbiotic structures and stages within a root system. Therefore tools allowing the analysis of molecular changes at a cellular level are required to reveal insight into arbuscular mycorrhizal (AM) symbiosis development and functioning.

Results: Here we describe the analysis of metabolite pools in arbuscule-containing cells, which are the site of nutrient transfer between AM fungus and host plant. Laser capture microdissection (LCM) combined with gas chromatography mass spectrometry (GC-EI/TOF-MS) enabled the analysis of primary metabolite levels,which might be of plant or fungal origin, within these cells.

Conclusions: High levels of the amino acids, aspartate, asparagine, glutamate, and glutamine, were observed in arbuscule-containing cells. Elevated amounts of sucrose and the steady-state of hexose levels indicated a direct assimilation of monosaccharides by the fungal partner.

No MeSH data available.