Hong-Fei Chen

Proteomic and phosphoproteomic determination of ABA's effects on grain-filling of Oryza sativa L. inferior spikelets

Cultivars of rice (Oryza sativa L.), especially the large-spikelet-type, often fail to achieve the high yield potential due to poor grain-filling of their inferior (late-flowering) spikelets. The superior (early-flowering) spikelets normally contain more abscisic acid (ABA) than the inferior spikelets. It was speculated that ABA might play a pivotal role in the grain-filling of inferior spikelets. To understand the molecular regulation involved in this process, we employed the 2-D gel-based comparative proteomic and phosphoproteomic analyses to search for differentially expressed proteins in the inferior spikelets under exogenous ABA treatment. A total of 111 significantly differential proteins and 31 phosphoproteins were found in the inferior spikelets after treatment. Among them, 100 proteins and 23 phosphoproteins were identified by using MALDI-TOF/TOF MS. In addition, the gene expression patterns of the inferior spikelets were confirmed with RT-PCR. These differentially expressed proteins are active in defense response, carbohydrate, protein, amino acid, energy and secondary metabolisms, as well as cell development and photosynthesis. The results suggest that the grain-filling of rice inferior spikelets is regulated by ABA through some proteins and phosphoproteins participating in carbon, nitrogen and energy metabolisms.

Differential Expression and Function Analysis of Proteins in Flag Leaves of Rice During Grain Filling

Abstract To investigate the molecular mechanism of metabolism in rice ( Oryza sativa L.) source organs, protein expression pattern in flag leaves of rice cultivar Jinhui 809, a cultivar with large panicle and high seed-setting percentage, was analyzed during grain-filling period. Using two-dimensional electrophoresis and mass spectrometry, 17 protein spots with differential expression were detected. Among them, 3 protein spots had the most expression amount from early to medium stage, 9 protein spots for middle–late stages, and 4 protein spots for late stage. In addition, 1 protein spot had two-expression peaks at early and late stages of grain filling. Twelve differential proteins were identified with MALDI-TOF/MS analysis and database searching. They were involved in biosynthesis and degradation, carbohydrate transportation, antioxidation reaction, hormone metabolism, cytoskeleton construction, and tissue maturation. Ribose/galactose/methyl galactoside importing ATP-binding protein 1 participated in photosynthate transportation from leaf to grain in early and medium stages. Auxin-responsive protein IAA27 influenced photosynthate transportation through regulating the activity of ATPase. N-acetyl-glutamate semialdehyde dehydrogenase delayed leaf senescence through regulating polyamine metabolism in late stage. In addition, glutathione S -transferase and superoxide dismutase played an important role in plant detoxification and fighting against active oxygen damage at late stage of grain filling.

Analysis of Differential Expression of Proteins in Rice Leaf Sheath during Grain Filling

Abstract To reveal the molecular mechanism of metabolism in rice ( Oryza sativa L.) leaf sheath, differential expression of proteome in rice leaf sheath was investigated with a large panicles and high grain-setting rate cultivar, i.e., Jinhui 809. Through the method of differential proteomics, 23 proteins were detected, and 11 proteins were identified to be the functional proteins. These functional proteins were classified into 6 categories according to their dynamic patterns of expression during grain-filling stage. The first class, such as oxaloacetate decarboxylase alpha subunit, showed down-regulation during grain filling. The second class, which included ribulose bisphosphate carboxylase small chain and ADP-ribosylation factor 1, exhibited up-regulation at first and then down-regulation. The third class, such as auxin response factor, zinc finger, C3HC4 type family protein, vacuolar proton-ATPase, and ribulose bisphosphate carboxylase/oxygenase activase, was down-regulation first and then up-regulation and down-regulation alternatively. The fourth class included Rubisco binding-protein alpha subunit, showing an expression pattern of up and down-regulation alternatively twice during grain filling. The fifth class performed a decreasing at first and then gradually increasing expression pattern. This class included class II metallothionein-like protein 1A and geranylgeranyl diphosphate synthase. The sixth class was protein kinase family protein performing a gradually increasing expression pattern. These 6 classes of proteins were involved in photosynthesis, hormone regulation, substance transportation, resistant response to plant senescence, and cell signal transduction in leaf sheath, respectively. They regulated the transition of sink to source together.

Effects of Nitrogen Management on Protein Expression of Flag Leaves at Grain-Filling Stage in Large Panicle Rice

To explain the metabolic mechanism of leaf during grain filling of rice (Oryza sativa L.) in response to nitrogen application, the differential expression patterns of leaf proteins were investigated at different time points of grain filling using two-dimensional electrophoresis (2-DE) technique. Thirty-two differentially expressed proteins were detected including 27 up-regulated and 5 down-regulated proteins in response to increased nitrogen application at grain-filling stage. These proteins were classified into 5 main categories according to their deduced functions, which were involved in photosynthesis (12), response to adversity resistance (5), hormone synthesis and signal transduction (5), cell growth and differentiation (5), and unknown functions (5). The photosynthesis-related features, together with adversity-defense-related indicators, were then determined to reveal the regulation of nitrogen to rice leaf. As appropriate increase of nitrogen application at late growth stage, the adversity-defense ability of leaf was strengthened during grain filling by delaying degradation of chlorophyll and soluble protein, prolonging photosynthesis, activating superoxide dismutase, peroxidase, and catalase, and depressing lipid peroxidation. These physiological changes verified the result of proteomics. Therefore, appropriate increased nitrogen supply at late growth stage is favorable to leaf metabolisms during grain filling in rice.

Differential proteomic analysis of rice seedlings reveals the advantage of dry-raising nursery practices

Dry-raising rice seedlings in nurseries is a key technique in high-yield rice cultivation. The present study of morphological and physiological indexes showed dry-raised seedlings (DRS) had a shorter stature, more developed root systems, and significantly higher soluble sugar, starch, and N content than moist-raised seedlings (MRS), resulting in significantly increased grain yield. Compared to the MRS techniques, the dry-raised measures induced higher levels of abscisic acid (ABA), gibberellins (GA3), and indole-3-acetic acid (IAA) in leaves and roots of seedlings. We then utilized tandem mass tags (TMT) quantitative proteomics technology to analyze the mechanism by which rice exposed to the appropriate drought stress (dry-raised measures) during the seedling stage develop differently. Through mass spectrometry, we identified 281 significantly expressed proteins in roots and 268 in leaves. The differentially expressed proteins were then divided into 23 categories based on MapMan ontology. In addition, the hormonal-related protein expression patterns of DRS were confirmed with RT-PCR at the transcript level. On the basis of these findings, we proposed that appropriate drought stress during the rice seedling stage can change the expression of key proteins involved in nitrogen uptake and translocation, hormone synthesis, photosynthesis, and CHO metabolic processes, thus regulating rice seedling growth. In this process, the differentially expressed key proteins, such as the 14-3-3 protein, GTP-binding protein, and calcium, play important roles in transduction of signals regarding soil drought, and the upregulated heat shock protein, glutathione S-transferases, and peroxidases function in enhancing the stress tolerance of the seedlings under dry-raising nursery conditions. This study established the high yielding mechanism of dry-raised cultivates methods during seedling stage at the protein expression level.