Creating a zero amylose barley with high soluble sugar content by genome editing
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Starch synthase
Wild type
Starch synthase
Branching (polymer chemistry)
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In order to provide reference to genetic improvement of wheat noodle quality,the activities of starch biosynthetic enzymes at grain-filling stage were investigated in grains of 4 wheat cultivars carrying all 3 Wx proteins subunits,which controlling the synthesis of amylose,starch physiochemical properties and noodle quality.Results showed that the activity of granule-bound starch synthase(GBSS) in developing endosperm from Jimai19 and Jimai20 was significantly lower than that of Lumai14 and Lumai21,while the activities of other key enzymes involved in amylopectin biosynthesis were similar,such as ADP-glucose pyrophosphorylase(AGPase),soluble starch synthase(SSS),and starch branching enzyme(SBE).Therefore,Jimai19 and Jimai20 exhibited lower amylose content in grain,and higher flour swelling power,pasting peak viscosity and breakdown,as well as better noodle quality.Activities of starch biosynthetic enzymes were correlated significantly with accumulating rate of amylose and amylopectin during grain filling,which suggested wheat cultivars were different in starch biosynthesis and processing quality mainly due to genotype.The activities of enzymes involved in starch biosynthesis were correlated significantly in developing endosperm,which indicated that they interacted in starch biosynthesis,the same was as accumulation of amylose and amylopectin.It is possible to breed wheat cultivars for preferred noodle quality from wild common wheat without Wx protein deficiency,and modification in activity or expression of one and several enzymes might not improve starch properties greatly.
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Two different wheat varieties were used to study the dynamic changes of the accumulations of amylose and amylopectin,starch accumulation rate and the activity changes of key enzymes involved in starch biosynthesis in the kernel during grain filling.The results showed that the beginning biosynthesis of amylopectin was earlier than that of amylose,based on the result of amylose appeared at 7 days after anthesis in non-waxy wheat,meanwhile,amylopectin appeared before 7 days after anthesis and the waxy wheat varieties only had amylopectin.The changes of amylose and amylopectin accumulating rates of both waxy and non-waxy wheat varieties showed a single peak curve.The accumulation courses of both amylopectin and amylose were well fitted to the logistic equation by relating amylopectin and amylose contents against days post anthesis.The simulation parameters revealed that the higher contents of amylopectin and amylose resulted from greater accumulation rate and mean accumulation rate,but accumulation duration probably played a less important role for them.The results indicated that the activity changes of the adenosine diphosphorate glucose pyrophrylase(AGPP),soluble starch synthase(SSS),granule-bound starch synthase(GBSS) and starch branching enzyme(SBE) were all in the pattern of a single-peak curve,and peaked at 20～25 days after anthesis.The accumulation rate of amylose was significantly or highly significantly correlated with the activities of AGPP,SSS,GBSS and SBE,the accumulation rate of amylopectin was significantly correlated with the activity of SSS,and the accumulation rate of starch was significantly or highly significantly correlated with the activities of AGPP and SSS.
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Starch synthase (SS) IIIa has the second highest activity of the total soluble SS activity in developing rice endosperm. Branching enzyme (BE) IIb is the major BE isozyme, and is strongly expressed in developing rice endosperm. A mutant (ss3a/be2b) was generated from wild-type japonica rice which lacks SSIIa activity. The seed weight of ss3a/be2b was 74-94% of that of the wild type, whereas the be2b seed weight was 59-73% of that of the wild type. There were significantly fewer amylopectin short chains [degree of polymerization (DP) ≤13] in ss3a/be2b compared with the wild type. In contrast, the amount of long chains (DP ≥25) connecting clusters of amylopectin in ss3a/be2b was higher than in the wild type and lower than in be2b. The apparent amylose content of ss3a/be2b was 45%, which was >1.5 times greater than that of either ss3a or be2b. Both SSIIIa and BEIIb deficiencies led to higher activity of ADP-glucose pyrophosphorylase (AGPase) and granule-bound starch synthase I (GBSSI), which partly explains the high amylose content in the ss3a/be2b endosperm. The percentage apparent amylose content of ss3a and ss3a/be2b at 10 days after flowering (DAF) was higher than that of the wild type and be2b. At 20 DAF, amylopectin biosynthesis in be2b and ss3a/be2b was not observed, whereas amylose biosynthesis in these lines was accelerated at 30 DAF. These data suggest that the high amylose content in the ss3a/be2b mutant results from higher amylose biosynthesis at two stages, up to 20 DAF and from 30 DAF to maturity.
Starch synthase
Wild type
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The amylose content of wheat grain is significantly correlated with amount of waxy protein subunit.However,there is a great difference in amylose content among different wheat cultivars which all have three waxy genes.In order to investigate the activity changes of enzymes involved in starch biosynthesis,the characteristics of starch accumulation in grains,and the relationship between them, two wheat(Triticum aestivum L.) cultivars,JM20(lower amylose content in grains) and LM21(higher amylose content in grains) were used in the study,which both carry three Waxy protein subunits.The results indicated that the activities of sucrose synthase(SS),adenosine diphosphorate glucose pyrophrylase(AGPP),soluble starch synthase(SSS),granule-bound starch synthase(GBSS) changed in the pattern of a single-peak curve during grain filling.The activities of SS,AGPP,GBSS,SSS and SBE in LM21,which had higher starch content,were higher than those in JM20.The accumulation rate of amylopectin was significantly or highly significantly correlated with the activities of SS,AGPP,SSS and SBE, and the accumulation rate of amylose was highly significantly correlated with the activities of SS,AGPP and GBSS.The accumulation courses of both amylopectin and amylose were well fitted to the logistic equation by relating amylopectin and amylose contents against days post anthesis.The simulation parameters revealed that the higher contents of amylopectin and amylose resulted from earlier initiating accumulation time and greater accumulation rate,but accumulation duration probably played a less important role for them.The amylose content in grains was mainly determined by GBSS activity as well as the activities of SS and AGPP.The change of GBSS activity reflected expression level of waxy gene was closely associated with the accumulation rate of amylose,indicating that amylose content in grains was determined by GBSS activity,especially at late grain filling stage.The difference in expression of waxy gene might be a major reason for the large difference in amylose contents between JM20 and LM21.
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【Objective】 This study was to elucidate the difference in starch physiochemical properties among wheat cultivars differing in Wx protein combinations through the comparison of starch accumulation and activities of starch biosynthetic enzymes.【Method】 The accumulation of amylose and amylopectin in developing endosperm from 14 wheat cultivars with six types of Wx protein combinations were investigated,together with activities of four key enzymes involved in starch biosynthesis such as ADP-glucose pyrophosphorylase(AGPase),soluble starch synthase(SSS),granule bound starch synthase(GBSS),and starch branching enzyme(SBE).【Result】 The number and type of Wx protein had a significant influence on activities of starch biosynthetic enzymes and the accumulation of amylose and amylopectin in developing endosperm.The activities of starch biosynthetic enzymes increased with the number of Wx protein,especially for GBSS,together with the amylose content in grain.The Wx-B1 protein deficiency induced the lowest GBSS activity and amylose content in grain,then was the Wx-D1 protein,and the effect of Wx-A1 protein deficiency was the least.The activities of four starch biosynthetic enzymes correlated to each other significantly,and also correlated to the number of Wx protein,amylose and amylopectin content in grain,which suggested the biosynthesis of amylose and amylopectin in developing endosperm were interacted and synchronous.【Conclusion】 The Wx allele is one of the most determinant factors for amylose content for it can control the activity of GBSS through the number and type of Wx protein and affect other starch biosynthetic enzymes indirectly,then the synthesis of amylose and amylopectin,which result in different starch physiochemical properties and noodle quality finally.
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Four rice varieties with different amylose contents were used to study the pattern of grain starch's accumulation and the activity of related enzymes,and the correlation between the pattern and activity was analyzed.The results showed that accumulation rate of amylase and amylopectin,granule bound starch synthase( GBSS) and soluble starch synthase( SSS) changed in the pattern of a single peak curve during grain filling. The growth of amylose content reached to the maximum in 5—15 days after flowering in all rice varieties,and was highest in Feiyou188,lowest in Chengnuo88. The content of amylopectin increased significantly in 5—20 days after flowering( P0. 01) and was highest in B6you4761. The accumulation courses of both amylopectin and amylose were well fitted to the logistic equation. The simulation of accumulation process discover that the maximum accumulation rate appears with higher maximum accumulation amount of amylose; and the accumulation amount of amylopectin is positive correlated with the an earlier appearance of the maximum accumulation rate and a longer active accumulation days.The accumulation amount and rate of amylose,GBSS activity in Feiyou88 with a high AC content were significantly higher than other varieties. The amylopectin accumulation and accumulation rate in Chengnuo 88 reached to the highest within 15 days after anthesis,but its SSS activity in grain was low. The 1000-grain weight of Feiyou88 was higher than that of other varieties during end filling stage. Correlation analysis indicates that there is a positive correlation between accumulation rate of grain starch and the activity of GBSS and SSS in all rice varieties,which illustrates that activity of GBSS and SSS affect the accumulation of grain starch.
Starch synthase
Anthesis
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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.
Starch synthase
Granule (geology)
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