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. 2019 Jan 16;20(2):362.
doi: 10.3390/ijms20020362.

Combined Proteomic and Metabolomic Profiling of the Arabidopsis thaliana vps29 Mutant Reveals Pleiotropic Functions of the Retromer in Seed Development

Thomas C Durand  1 , Gwendal Cueff  2 , Béatrice Godin  3 , Benoît Valot  4 , Gilles Clément  5 , Thierry Gaude  6 , Loïc Rajjou  7
Affiliations

Combined Proteomic and Metabolomic Profiling of the Arabidopsis thaliana vps29 Mutant Reveals Pleiotropic Functions of the Retromer in Seed Development

Thomas C Durand et al. Int J Mol Sci. .

Abstract

The retromer is a multiprotein complex conserved from yeast to humans, which is involved in intracellular protein trafficking and protein recycling. Selection of cargo proteins transported by the retromer depends on the core retromer subunit composed of the three vacuolar protein sorting (VPS) proteins, namely VPS26, VPS29, and VPS35. To gain a better knowledge of the importance of the plant retromer in protein sorting, we carried out a comparative proteomic and metabolomic analysis of Arabidopsis thaliana seeds from the wild-type and the null-retromer mutant vps29. Here, we report that the retromer mutant displays major alterations in the maturation of seed storage proteins and synthesis of lipid reserves, which are accompanied by severely impaired seed vigor and longevity. We also show that the lack of retromer components is counterbalanced by an increase in proteins involved in intracellular trafficking, notably members of the Ras-related proteins in brain (RAB) family proteins. Our study suggests that loss of the retromer stimulates energy metabolism, affects many metabolic pathways, including that of cell wall biogenesis, and triggers an osmotic stress response, underlining the importance of retromer function in seed biology.

Keywords: Arabidopsis; germination; longevity; metabolomics; proteomics; retromer; seed; vacuolar protein sorting 29.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Impact of the loss of retromer function on seed germination and longevity. (a) Germination curve of Col-0 and vps29 Arabidopsis seeds after cold stratification treatment (5 days at 4 °C in the dark) and subsequent transferral to continuous light at 25 °C. Seed germination (considered as radicle protrusion through the seed coat and the endosperm) was scored daily. White circles represent the germination means for Col-0 seeds. Black circles represent the germination means for vps29 seeds. The bars represent the standard deviation (±SD) of the mean of three independent experiments. (b) Influence of controlled deterioration treatment (CDT) on Arabidopsis seed germination. Col-0 and vps29 seeds were submitted to CDT for different periods of time (0, 1, 3, and 5 days). The graph shows the maximum percentage of germination (Gmax) of deteriorated seeds. It is a representative experiment carried out three times in triplicate. The bars represent the standard deviation (±SD) of the mean of three independent experiments. Dashed lines represent the polynomial curve-fit related to these following equations: Col-0, y = −2.3054x2 + 5.4464x + 90.901 with R2 = 0.99037; vps29, y = 1.5537x2 − 26.453x + 107.67 with R2 = 0.98204.
Figure 2
Figure 2
Influence of the retromer VPS29 mutation on Arabidopsis dry seed proteome. An equal amount (150 µg) of total protein extracts was loaded on each gel. (a) A representative silver nitrate-stained 2DE gel of total soluble proteins from Col-0 (Arabidopsis, accession Columbia-0; wild-type) dry mature seeds. (b) A representative silver nitrate-stained 2DE gel of total soluble proteins from vps29 dry mature seeds. The rectangles highlight the protein clusters of 12S storage protein precursors (p), 12S alpha-cruciferins (α), and 12S beta-cruciferins (β). MW, molecular weight; pI, isoelectric point.
Figure 3
Figure 3
Gene ontology (GO) term enrichment analysis of the selected 442 differentially abundant proteins in vps29 dry seeds in comparison with Col-0 seeds. (a) Overrepresented GO terms for 115 proteins that were up-accumulated in Col-0 dry seeds. (b) Overrepresented GO terms for 327 proteins that were up-accumulated in vps29 dry seeds. Data from PANTHER overrepresentation test (http://www.geneontology.org); Arabidopsis thaliana GO database released 1 June 2018. Black bars: observed proteins; grey bars: expected result from the reference Arabidopsis genome.
Figure 4
Figure 4
The energy metabolism is altered in vps29 Arabidopsis thaliana seeds. Numbers preceeded by an upward or downward arrow indicate the vps29/Col-0 positive (in red) or negative (in blue) abundance ratio of the enzymes (see Table S2) or metabolites (see Table 1, Table 2, and Table S4), respectively.
Figure 5
Figure 5
Schematic diagram illustrating the main effects related to vps29 mutation in Arabidopsis thaliana seeds highlighted by proteomics and metabolomics.
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