The Proteome of Seed Development in the Model Legume Lotus japonicus      

Abstract

We have characterized the development of seeds in the model legume Lotus japonicus. Like soybean (Glycine max) and pea (Pisum sativum), Lotus develops straight seed pods and each pod contains approximately 20 seeds that reach maturity within 40 days. Histological sections show the characteristic three developmental phases of legume seeds and the presence of embryo, endosperm, and seed coat in desiccated seeds. Furthermore, protein, oil, starch, phytic acid, and ash contents were determined, and this indicates that the composition of mature Lotus seed is more similar to soybean than to pea. In a first attempt to determine the seed proteome, both a two-dimensional polyacrylamide gel electrophoresis approach and a gel-based liquid chromatography-mass spectrometry approach were used. Globulins were analyzed by two-dimensional polyacrylamide gel electrophoresis, and five legumins, LLP1 to LLP5, and two convicilins, LCP1 and LCP2, were identified by matrix-assisted laser desorption ionization quadrupole/time-of-flight mass spectrometry. For two distinct developmental phases, seed filling and desiccation, a gel-based liquid chromatography-mass spectrometry approach was used, and 665 and 181 unique proteins corresponding to gene accession numbers were identified for the two phases, respectively. All of the proteome data, including the experimental data and mass spectrometry spectra peaks, were collected in a database that is available to the scientific community via a Web interface (http://www.cbs.dtu.dk/cgi-bin/lotus/db.cgi). This database establishes the basis for relating physiology, biochemistry, and regulation of seed development in Lotus. Together with a new Web interface (http:// bioinfoserver.rsbs.anu.edu.au/utils/PathExpress4legumes/) collecting all protein identifications for Lotus, Medicago, and soybean seed proteomes, this database is a valuable resource for comparative seed proteomics and pathway analysis within and beyond the legume family.

Post-Genomics Studies of Developmental Processes in Legume Seeds

“…More comprehensive sequence information also permits the analysis of various posttranslational modifications that occur and increase the number of polypeptides arising from a single sequence. A reconstruction of posttranslational modifications leading to the deposition of mature globulins was elegantly demonstrated by Dam et al (2009) in L. japonicus and by Bourgeois et al (2009) in pea (Pisum sativum). Dam et al (2009) also compared the protein compositions of green and mature seeds using gel filtration liquid chromatography/tandem mass spectrometry (MS/MS) and identified 920 proteins from green and 264 proteins from mature seeds.…”

“…For the entire seed, the citations refer to post-genomics experiments carried out in various legume species at the level of the transcriptome (Firnhaber et al, 2005;Buitink et al, 2006;Gallardo et al, 2007;Benedito et al, 2008;Verdier et al, 2008), metabolome (Vigeolas et al, 2008), ionome (Sankaran et al, 2009), and proteome. At the level of the proteome, some citations refer to the mature seed (Watson et al, 2003;Magni et al, 2007;Bourgeois et al, 2009;Krishnan et al, 2009;Natarajan et al, 2009), to the kinetics of seed development (Hajduch et al, 2005;Gallardo et al, 2007;Agrawal et al, 2008;Dam et al, 2009), and to a functional study for identifying targets of thioredoxin (Alkhalfioui et al, 2007). For the separated seed tissues, the citations refer to transcriptome and proteome analyses in soybean (Le et al, 2007) and M. truncatula (Boudet et al, 2006;Zhang et al, 2006;Gallardo et al, 2007).…”

“…Suc synthase, starch synthase, chlorophyll a/b-binding protein, oxygen-evolving enhancer protein; Gallardo et al, 2003). A b-amylase involved in the irreversible pathway of starch breakdown was identified by Dam et al (2009) in developing L. japonicus seeds that may play a role in starch remobilization with the seed.…”

Two-dimensional gel electrophoretic protein profile analysis during seed development of Ocotea catharinensis: a recalcitrant seed species

“…Over latter years, several studies have focused on characterizing protein dynamics during plant development, as well as the associated genomes and transcriptomes involved (Roberts 2002;Heazlewood and Millar 2003;Chen and Harmon 2006;Gallardo et al 2007;Dam et al 2009). It has been demonstrated that proteomic analysis can generate relevant information towards elucidating the identity of those molecular mechanisms involved in developmental processes.…”

Seed germination in Ormosia macrocalyx , an endangered tropical forest tree

“…Ormosia macrocalyx seeds obtained from completely open fruits that were exposed to the environment, presented an important delay in germination initiation (almost 4 days) relative to those from closed or semi-open fruits. Taking into consideration the notion proposed by Dam et al (2009) that fruit and seeds pass through phases of histological differentiation, seed filling and desiccation, our results suggest that dry but still closed O. macrocalyx fruits contain seeds with mature and viable embryos. Once the fruits are open (dehiscence), their seeds become subject to desiccation, which promotes dispersion, with the consequent development of seed dormancy (Silva et al 2014).…”