Proteomics of Arabidopsis Seed Germination. A Comparative Study of Wild-Type and Gibberellin-Deficient Seeds

Abstract

We examined the role of gibberellins (GAs) in germination of Arabidopsis seeds by a proteomic approach. For that purpose, we used two systems. The first system consisted of seeds of the GA-deficient ga1 mutant, and the second corresponded to wild-type seeds incubated in paclobutrazol, a specific GA biosynthesis inhibitor. With both systems, radicle protrusion was strictly dependent on exogenous GAs. The proteomic analysis indicated that GAs do not participate in many processes involved in germination sensu stricto (prior to radicle protrusion), as, for example, the initial mobilization of seed protein and lipid reserves. Out of 46 protein changes detected during germination sensu stricto (1 d of incubation on water), only one, corresponding to the cytoskeleton component ␣-2,4 tubulin, appeared to depend on the action of GAs. An increase in this protein spot was noted for the wild-type seeds but not for the ga1 seeds incubated for 1 d on water. In contrast, GAs appeared to be involved, directly or indirectly, in controlling the abundance of several proteins associated with radicle protrusion. This is the case for two isoforms of S-adenosyl-methionine (Ado-Met) synthetase, which catalyzes the formation of Ado-Met from Met and ATP. Owing to the housekeeping functions of Ado-Met, this event is presumably required for germination and seedling establishment, and might represent a major metabolic control of seedling establishment. GAs can also play a role in controlling the abundance of a ␤-glucosidase, which might be involved in the embryo cell wall loosening needed for cell elongation and radicle extension.Maturation drying is the normal terminal event in the vast majority of seeds, after which they pass into a metabolically quiescent state where they may remain for many years and still retain their viability (Hoekstra et al., 2001). Upon hydration under suitable conditions, the seed, if not dormant, reactivates its metabolism and commences germination, giving rise to a new plant.Seed germination can be divided into three phases, imbibition, increased metabolic activity, and initiation of growth, which loosely parallel the triphasic water uptake of dry mature seeds. Morphologically, initiation of growth corresponds to radicle emergence; subsequent growth is generally defined as seedling growth. By definition, germination sensu stricto incorporates those events that start with the uptake of water by the quiescent dry seed and terminate with the protrusion of the radicle and the elongation of the embryonic axis (Bewley, 1997). From physiological studies on a wide variety of species, including a number of mutants, it appears that gibberellins (GAs) play a key role in late stages of seed germination Karssen, 1988, 1992;Karssen et al., 1989;Hilhorst and Toorop, 1997; Yamaguchi et al., 1998; Richards et al., 2001). Thus, in plant species such as Arabidopsis and tomato (Lycopersicon esculentum), the strong alleles of GA-deficient mutants are unable to complete germination without exogenous GAs (Koornneef and van der Veen,...

Changes in the root proteome of Triticosecale grains germinating under osmotic stress

“…The extraction of proteins was carried out according to the procedure proposed by Gallardo et al (2002). About 1 g of frozen root tissue was carefully pulverized in mortars cooled with liquid nitrogen and the material transferred to Eppendorf tubes with a suitable amount (20 ll/mg dry weight of tissue) of lysis buffer containing: 7 M urea; 2 M thiourea; 4 % CHAPS; 40 mM dithiothreitol; 2 % ampholytes pH 3-11; 60 U/ml DNase I; 5.8 Kunitz/ml RNase A; 1 pill/10 ml protease inhibitor Cocktail Tablets cOmplete Mini (Roche).…”

Enzymatic characterization of germination-specific cysteine protease-1 expressed transiently in cotyledons during the early phase of germination

“…Furthermore, storage of the enzyme at À30 C for 1 month resulted in almost complete loss of activity, and the absence of a 26 kDa band on SDS PAGE. This instability and transient expression of the enzyme could account for the enzyme failing to be identified at the protein level, even by proteomics (27) or highly sensitive activity profiling analysis of cysteine proteases (15) of A. thaliana. Except for Brassicaceae, DRCP26 is highly homologous with putative cysteine proteases from grapes (Vitis vinifera, UniProt ID: F6H4F8, amino acid identity: 73%), kidney bean (Phaseolus vulgaris, UniProt ID: O24321, 70%), soybean (Glycine Max, UniProt ID: C6TFM1, 69%), upland cotton (Gossypium hirsutum, UniProt ID: G8H1L9, 69%), vetch (Vicia sativa, UniProt ID: Q41696, 68%) (28) and chickpea (Cicer arietinum, UniProt ID: Q39452, 66%) (29).…”

Effects of carbon and nitrogen content on seed germination of calabrian pine (Pinus brutia) populations

“…The effects of germination on the chemical composition and biochemical constituents of seeds vary greatly with the plant species, seed varieties or cultivars and germination conditions (Gallardo et al 2002). Proteins are necessary for seed germination, and they accumulate after ripening under seed-drying conditions, resulting in the release of dormancy (Gallardo et al 2002, Chibani et al 2006. Among the most important parameters that control the process of seed dormancy are changes associated with protein and hormones (Finch-Savage and Leubner-Metzger 2006, Finkelstein et al 2008, Graeber et al 2012, Miransari and Smith 2014.…”

“…They also determined that there was a relationship of R 2 = 0.69 between the seed germination rate and seed carbon concentrations and a relationship of R 2 = 0.11 between the seed germination rate and seed nitrogen concentrations. The effects of germination on the chemical composition and biochemical constituents of seeds vary greatly with the plant species, seed varieties or cultivars and germination conditions (Gallardo et al 2002). Proteins are necessary for seed germination, and they accumulate after ripening under seed-drying conditions, resulting in the release of dormancy (Gallardo et al 2002, Chibani et al 2006.…”