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Biosynthesis and Regulation of Wheat Amylose and Amylopectin from Proteomic and Phosphoproteomic Characterization of Granule-binding Proteins

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ABSTRACT

Waxy starch has an important influence on the qualities of breads. Generally, grain weight and yield in waxy wheat (Triticum aestivum L.) are significantly lower than in bread wheat. In this study, we performed the first proteomic and phosphoproteomic analyses of starch granule-binding proteins by comparing the waxy wheat cultivar Shannong 119 and the bread wheat cultivar Nongda 5181. These results indicate that reduced amylose content does not affect amylopectin synthesis, but it causes significant reduction of total starch biosynthesis, grain size, weight and grain yield. Two-dimensional differential in-gel electrophoresis identified 40 differentially expressed protein (DEP) spots in waxy and non-waxy wheats, which belonged mainly to starch synthase (SS) I, SS IIa and granule-bound SS I. Most DEPs involved in amylopectin synthesis showed a similar expression pattern during grain development, suggesting relatively independent amylose and amylopectin synthesis pathways. Phosphoproteome analysis of starch granule-binding proteins, using TiO2 microcolumns and LC-MS/MS, showed that the total number of phosphoproteins and their phosphorylation levels in ND5181 were significantly higher than in SN119, but proteins controlling amylopectin synthesis had similar phosphorylation levels. Our results revealed the lack of amylose did not affect the expression and phosphorylation of the starch granule-binding proteins involved in amylopectin biosynthesis.

No MeSH data available.


Quantitative real-time PCR (qRT-PCR) analysis of 6 key genes related to starch synthesis in developing seeds.
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f6: Quantitative real-time PCR (qRT-PCR) analysis of 6 key genes related to starch synthesis in developing seeds.

Mentions: The dynamic expression profiles of 16 main starch synthesis-related genes during 10 grain developmental stages in the waxy wheat SN119 and non-waxy wheat ND5181 were analyzed by qRT-PCR (Fig. 6). In general, starch synthesis-related genes in SN119 and ND5181 displayed similar expression patterns during grain development. Most of them showed an up-down expression trend and had higher expression levels during the middle stages of grain development, but ISA III and AGPS I displayed down-up and down expression trends, respectively. GBSS I exhibited an up-down expression pattern in the bread wheat ND5181, but was barely expressed in the waxy wheat SN119 (Fig. 6A). Four genes (phosphoglucomutase (PGM), Phosphoglucoisomerase (PGI), ADPglucose pyrophosphorylase large subunits (AGPL I) and ADPglucose pyrophosphorylase small subunits (AGPS II) involved in the early stages of starch biosynthesis were expressed early and their expression peaked at about 12 DPA (Fig. 6B,C). Five genes (SS IIa, SS III, SBE I, SBE IIa and SBE IIb), which control the synthesis of longer chains (B1/B2-chains) were highly expressed at 12–15 DPA, while SS I and SBE I, which control the synthesis of shorter chains (A-chains), were expressed abundantly at 17–20 DPA (Fig. 6D,E). DBEs mainly showed an up-down expression pattern, with the exception of ISA III (Fig. 6F). Dynamic transcriptional expression profiling of starch biosynthesis-related genes showed similar transcriptional expression profiles in the two cultivars. The transcriptional and protein expression patterns of SS I, SS IIa and GBSS I exhibited up-down trends in both SN119 and ND5181.


Biosynthesis and Regulation of Wheat Amylose and Amylopectin from Proteomic and Phosphoproteomic Characterization of Granule-binding Proteins
Quantitative real-time PCR (qRT-PCR) analysis of 6 key genes related to starch synthesis in developing seeds.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5015113&req=5

f6: Quantitative real-time PCR (qRT-PCR) analysis of 6 key genes related to starch synthesis in developing seeds.
Mentions: The dynamic expression profiles of 16 main starch synthesis-related genes during 10 grain developmental stages in the waxy wheat SN119 and non-waxy wheat ND5181 were analyzed by qRT-PCR (Fig. 6). In general, starch synthesis-related genes in SN119 and ND5181 displayed similar expression patterns during grain development. Most of them showed an up-down expression trend and had higher expression levels during the middle stages of grain development, but ISA III and AGPS I displayed down-up and down expression trends, respectively. GBSS I exhibited an up-down expression pattern in the bread wheat ND5181, but was barely expressed in the waxy wheat SN119 (Fig. 6A). Four genes (phosphoglucomutase (PGM), Phosphoglucoisomerase (PGI), ADPglucose pyrophosphorylase large subunits (AGPL I) and ADPglucose pyrophosphorylase small subunits (AGPS II) involved in the early stages of starch biosynthesis were expressed early and their expression peaked at about 12 DPA (Fig. 6B,C). Five genes (SS IIa, SS III, SBE I, SBE IIa and SBE IIb), which control the synthesis of longer chains (B1/B2-chains) were highly expressed at 12–15 DPA, while SS I and SBE I, which control the synthesis of shorter chains (A-chains), were expressed abundantly at 17–20 DPA (Fig. 6D,E). DBEs mainly showed an up-down expression pattern, with the exception of ISA III (Fig. 6F). Dynamic transcriptional expression profiling of starch biosynthesis-related genes showed similar transcriptional expression profiles in the two cultivars. The transcriptional and protein expression patterns of SS I, SS IIa and GBSS I exhibited up-down trends in both SN119 and ND5181.

View Article: PubMed Central - PubMed

ABSTRACT

Waxy starch has an important influence on the qualities of breads. Generally, grain weight and yield in waxy wheat (Triticum aestivum L.) are significantly lower than in bread wheat. In this study, we performed the first proteomic and phosphoproteomic analyses of starch granule-binding proteins by comparing the waxy wheat cultivar Shannong 119 and the bread wheat cultivar Nongda 5181. These results indicate that reduced amylose content does not affect amylopectin synthesis, but it causes significant reduction of total starch biosynthesis, grain size, weight and grain yield. Two-dimensional differential in-gel electrophoresis identified 40 differentially expressed protein (DEP) spots in waxy and non-waxy wheats, which belonged mainly to starch synthase (SS) I, SS IIa and granule-bound SS I. Most DEPs involved in amylopectin synthesis showed a similar expression pattern during grain development, suggesting relatively independent amylose and amylopectin synthesis pathways. Phosphoproteome analysis of starch granule-binding proteins, using TiO2 microcolumns and LC-MS/MS, showed that the total number of phosphoproteins and their phosphorylation levels in ND5181 were significantly higher than in SN119, but proteins controlling amylopectin synthesis had similar phosphorylation levels. Our results revealed the lack of amylose did not affect the expression and phosphorylation of the starch granule-binding proteins involved in amylopectin biosynthesis.

No MeSH data available.