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Complementary Proteome and Transcriptome Profiling in Developing Grains of a Notched-Belly Rice Mutant Reveals Key Pathways Involved in Chalkiness Formation.

ABSTRACT: Rice grain chalkiness is a highly complex trait involved in multiple metabolic pathways and controlled by polygenes and growth conditions. To uncover novel aspects of chalkiness formation, we performed an integrated profiling of gene activity in the developing grains of a notched-belly rice mutant. Using exhaustive tandem mass spectrometry-based shotgun proteomics and whole-genome RNA sequencing to generate a nearly complete catalog of expressed mRNAs and proteins, we reliably identified 38,476 transcripts and 3,840 proteins. Comparison between the translucent part and chalky part of the notched-belly grains resulted in only a few differently express genes (240) and differently express proteins (363), thus making it possible to focus on 'core' genes or common pathways. Several novel key pathways were identified as of relevance to chalkiness formation, in particular the shift of C and N metabolism, the down-regulation of ribosomal proteins and the resulting low abundance of storage proteins especially the 13 kDa prolamin subunit, and the suppressed photosynthetic capacity in the pericarp of the chalky part. Further, genes and proteins as transporters for carbohydrates, amino acid/peptides, proteins, lipids and inorganic ions showed an increasing expression pattern in the chalky part of the notched-belly grains. Similarly, transcripts and proteins of receptors for auxin, ABA, ethylene and brassinosteroid were also up-regulated. In summary, this joint analysis of transcript and protein profiles provides a comprehensive reference map of gene activity regarding the physiological state in the chalky endosperm.

SUBMITTER: Lin Z

PROVIDER: S-EPMC5444571 | biostudies-literature | 2017 Mar

REPOSITORIES: biostudies-literature

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Complementary Proteome and Transcriptome Profiling in Developing Grains of a Notched-Belly Rice Mutant Reveals Key Pathways Involved in Chalkiness Formation.

Lin Zhaomiao Z Wang Zunxin Z Zhang Xincheng X Liu Zhenghui Z Li Ganghua G Wang Shaohua S Ding Yanfeng Y

Plant & cell physiology 20170301 3

Rice grain chalkiness is a highly complex trait involved in multiple metabolic pathways and controlled by polygenes and growth conditions. To uncover novel aspects of chalkiness formation, we performed an integrated profiling of gene activity in the developing grains of a notched-belly rice mutant. Using exhaustive tandem mass spectrometry-based shotgun proteomics and whole-genome RNA sequencing to generate a nearly complete catalog of expressed mRNAs and proteins, we reliably identified 38,476 ...[more]

PMID: 28158863

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Project description:Grain chalkiness is a key quality trait of the rice grain, whereas its underlying mechanism is still not thoroughly understood because of the complex genetic and environmental interactions. We identified a notched-belly (NB) mutant that has a notched-line on the belly of grains. The line dissects the endosperm into two distinct parts, the upper translucent part, and the bottom chalky part in the vicinity of the embryo. Using this mutant, our previous studies clued the negative influence of embryo on the biochemical makeup of the endosperm, suggesting the need for the in-depth study of the embryo effect on the metabolome of developing endosperm. This study continued to use the NB mutant to evolve a novel comparison method to clarify the role of embryo in the formation of a chalky endosperm. Grain samples of the wild-type (WT) and NB were harvested at 10, 20, and 30 days after fertilization (DAF), and then divided into subsamples of the embryo, the upper endosperm, and the bottom endosperm. Using non-targeted metabolomics and whole-genome RNA sequencing (RNA-seq), a nearly complete catalog of expressed metabolites and genes was generated. Results showed that the embryo impaired the storage of sucrose, amino acid, starch, and storage proteins in the bottom endosperm of NB by enhancing the expression of sugar, amino acids, and peptide transporters, and declining the expression of starch, prolamin, and glutelin synthesis-related genes. Importantly, the competitive advantage of the developing embryo in extracting the nutrients from the endosperm, transformed the bottom endosperm into an "exhaustive source" by diverting the carbon (C) and nitrogen (N) metabolism from synthetic storage to secondary pathways, resulting in impaired filling of the bottom endosperm and subsequently the formation of chalky tissue. In summary, this study reveals that embryo-induced metabolic shift in the endosperm is associated with the occurrence of grain chalkiness, which is of relevance to the development of high-quality rice by balancing the embryo-endosperm interaction.

Project description:BackgroundThe analyses of protein synthesis, accumulation and regulation during grain development in wheat are more complex because of its larger genome size compared to model plants such as Arabidopsis and rice. In this study, grains from two wheat cultivars Jimai 20 and Zhoumai 16 with different gluten quality properties were harvested at five development stages, and were used to displayed variable expression patterns of grain proteins.ResultsProteome characterization during grain development in Chinese bread wheat cultivars Jimai 20 and Zhoumai 16 with different quality properties was investigated by 2-DE and tandem MALDI-TOF/TOF-MS. Identification of 117 differentially accumulated protein spots representing 82 unique proteins and five main expression patterns enabled a chronological description of wheat grain formation. Significant proteome expression differences between the two cultivars were found; these included 14 protein spots that accumulated in both cultivars but with different patterns and 27 cultivar-different spots. Among the cultivar-different protein spots, 14 accumulated in higher abundance in Jimai 20 than in Zhoumai 16, and included NAD-dependent isocitrate dehydrogenase, triticin precursor, LMW-s glutenin subunit and replication factor C-like protein. These proteins are likely to be associated with superior gluten quality. In addition, some proteins such as class II chitinase and peroxidase 1 with isoforms in developing grains were shown to be phosphorylated by Pro-Q Diamond staining and phosphorprotein site prediction. Phosphorylation could have important roles in wheat grain development. qRT-PCR analysis demonstrated that transcriptional and translational expression patterns of many genes were significantly different.ConclusionsWheat grain proteins displayed variable expression patterns at different developmental stages and a considerable number of protein spots showed differential accumulation between two cultivars. Differences in seed storage proteins were considered to be related to different quality performance of the flour from these wheat cultivars. Some proteins with isoforms were phosphorylated, and this may reflect their importance in grain development. Our results provide new insights into proteome characterization during grain development in different wheat genotypes.

Project description:Global warming impairs grain filling in rice and reduces starch accumulation in the endosperm, leading to chalky-appearing grains, which damages their market value. We found previously that high temperature-induced expression of starch-lytic ?-amylases during ripening is crucial for grain chalkiness. Because the rice genome carries at least eight functional ?-amylase genes, identification of the ?-amylase(s) that contribute most strongly to the production of chalky grains could accelerate efficient breeding. To identify ?-amylase genes responsible for the production of chalky grains, we characterized the histological expression pattern of eight ?-amylase genes and the influences of their overexpression on grain appearance and carbohydrate components through a series of experiments with transgenic rice plants. The promoter activity of most ?-amylase genes was elevated to various extents at high temperature. Among them, the expression of Amy1A and Amy3C was induced in the internal, especially basal to dorsal, region of developing endosperm, whereas that of Amy3D was confined near the ventral aleurone. These regions coincided with the site of occurrence of chalkiness, which was in clear contrast to conventionally known expression patterns of the enzyme in the scutellum and aleurone during seed germination. Furthermore, overexpression of ?-amylase genes, except for Amy3E, in developing endosperm produced various degrees of chalky grains without heat exposure, whereas that of Amy3E yielded normal translucent grains, as was the case in the vector control, even though Amy3E-overexpressing grains contained enhanced ?-amylase activities. The weight of the chalky grains was decreased due to reduced amounts of starch, and microscopic observation of the chalky part of these grains revealed that their endosperm consisted of loosely packed round starch granules that had numerous pits on their surface, confirming the hydrolysis of the starch reserve by ?-amylases. Moreover, the chalky grains contained increased amounts of soluble sugars including maltooligosaccharides at the expense of starch. The integrated analyses proposed that expression of Amy1A, Amy3C, and Amy3D at the specific regions of the developing endosperm could generate the chalkiness. This finding provides the fundamental knowledge to narrow down the targets for the development of high temperature-tolerant premium rice.

Dataset Information

Complementary Proteome and Transcriptome Profiling in Developing Grains of a Notched-Belly Rice Mutant Reveals Key Pathways Involved in Chalkiness Formation.

Publications

Complementary Proteome and Transcriptome Profiling in Developing Grains of a Notched-Belly Rice Mutant Reveals Key Pathways Involved in Chalkiness Formation.

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