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

Abstract

The aim of the present work was to characterize changes in the protein profile throughout seed development in O. catharinensis, a recalcitrant species, by two-dimensional gel electrophoresis. Protein extraction was undertaken by using a thiourea/urea buffer, followed by a precipitation step with 10% TCA. Comparative analysis during seed development showed that a large number of proteins were exclusively detected in each developmental stage. The cotyledonary stage, which represents the transition phase between embryogenesis and the beginning of metabolism related to maturation, presents the highest number of stage-specific spots. Protein identification, through MS/MS analysis, resulted in the identification of proteins mainly related to oxidative metabolism and storage synthesis. These findings contribute to a better understanding of protein metabolism during seed development in recalcitrant seeds, besides providing information on established markers that could be useful in defining and improving somatic embryogenesis protocols, besides monitoring the development of somatic embryos in this species.Key words: 2-DE; comparative proteomics; embryo development; storage protein; Lauraceae resumo O objetivo deste trabalho foi a caracterização dos perfis protéicos expressos durante o desenvolvimento da semente de O. catharinensis, uma espécie recalcitrante, via eletroforese bidimensional. A extração de proteínas foi realizada utilizando tampão uréia/tiouréia seguido de uma etapa de precipitação com TCA 10%. A análise comparativa durante o desenvolvimento da semente mostrou um grande número de proteínas exclusivas em cada estádio do desenvolvimento. O estádio cotiledonar, que representa a fase de transição entre a embriogênese e o começo do metabolismo relacionado a maturação, apresentou o maior número de proteínas estádio-específicas. Proteínas relacionadas ao metabolismo oxidativo e a síntese de reservas foram identificadas via MS/ MS. Estes resultados contribuem para o melhor entendimento do metabolismo de proteínas durante o desenvolvimento de sementes recalcitrantes, além de prover informações no estabelecimento de parâmetros para protocolos de embriogênese somática. palavras-chave: 2-DE; desenvolvimento embrionário; Lauraceae; proteômica comparativa; proteínas de reserva Braz.

Protein profile of cotyledon, tegument, and embryonic axis of mature acorns from a non-orthodox plant species: Quercus ilex

“…Significant changes in spot abundance are shown on one-and three-dimensional space Planta results, it has been reported that the ascorbate system is the main route to oxidative metabolism during embryogenesis and seed filling (De Tulio and Arrigoni 2003). The presence of the proteins related to the oxidative system in our mature seed is in agreement with observations by Pukacka and Ratajczak (2007) and Dias et al (2010). These authors demonstrated that recalcitrant seeds, in the case of O. catharinensis, are characterized by high ascorbate content and intensely active ascorbate system enzymes, when compared with orthodox seeds, in which ascorbic acid contents and ascorbate peroxidase activity decrease significantly or totally disappear during maturation and dehydration phases (Pukacka and Ratajczak 2007).…”

“…Ocotea catharinensis, a recalcitrant seed, showed an accumulation of metabolic protein and especially a carbohydrate metabolism (Dias et al 2010), which are key enzymes in cell wall biosynthesis (Chen et al 2007). The large number of energy and metabolicrelated spots in our mature seeds can indicate an active metabolism at the maturation phase.…”

Comparative proteomic analysis of somatic embryo maturation in Carica papaya L.

“…It acts in the translocation of proteins through the endoplasmic reticulum and assists in the proper folding and maturation of newly synthesized proteins entering the organelle. Therefore, this protein has been implicated in seed maturation in several species, such as Cucurbita maxima [ 87 ], Ocotea catharinensis [ 88 ] and Pinus massoniana [ 85 ]. However, overexpression of this protein inhibits the accumulation of seed storage proteins in Oryza sativa endosperm cells [ 89 ].…”

Germination and Early Seedling Development in Quercus ilex Recalcitrant and Non-dormant Seeds: Targeted Transcriptional, Hormonal, and Sugar Analysis

“…In orthodox seeds, germination represents a switch from quiescence in mature seeds to a metabolism reactivation under a low oxygen concentration in germinating ones ( Logan et al, 2001 ). In contrast, in recalcitrant seeds, maturation and germination is continuous, with an active metabolism also after shedding ( Dias et al, 2010 ; Sghaier-Hammami et al, 2016 ). Figure 6 shows the variation in the transcript abundance of four genes representing metabolic activity, Gapdh , Nadh6 , Rbcl , and Fdh , throughout the germination and early post-germination stages.…”