| Cassava (Manihot esculenta Crantz) is a staple food crop in the world with high photosynthetic efficiency in leaves and high starch accumulation in the tuberous roots. Tuberous roots are the main storage for starch synthesis and accumulation in cassava. How to increase sugar (assimilation production of leaves) to transform into starch and accumulate efficiently in the tuberous roots has been the main goals for cassava breeders? Sugar cassava was native to tropical America Amazonia. This germplasm has high tuber yield with 45 t/ha. It had high soluble sugar content in roots, but lower dry matter content and starch content, which was the best material to study starch accumulation and glucose metabolism in cassava. In the present study, South China 5 (SC5) was used as a control, which was the main culivar in China with high starch content. The changes regarding sugar and starch metabolism in different developmental stages (formation, enlargement and maturity) of sugar cassava (CAS-36-0, CAS-36-12) introduced from Brazil were detected in a global protein level. The starch contents and the soluble sugar contents including sucrose, glucose and fructose at different developmental stages of tuberous roots in three genotypes were measured using HPLC. The expression levels of SPS, UGPase, AGPase and Alpha-amlyase (AMY) were determined using Western blot. In combination with proteomic technology, the molecular mechanism of sugar metabolism in three cassava genotypes was elaborated to provide theoretical support for selecting cassava varieties with high quality and high starch content. The main results wre as follows:1. Starch content and amylose content of sugar cassava were much lower than that of SC5 at three developmental stages in tuberous root. The maximum in sugar cassava was in enlargement period, while SC5 was in mature period.2. The maximum of sucrose content in sugar cassava was in the formation period and minimum was in mature period. However, the maximum of sucrose content in SC5 was in mature period. Sucrose contents of sugar cassava were much lower than SC5 at mature period. While glucose and fructose contents of sugar cassava were much higher than SC5 at three developmental stages.3. Analysis of expression levels of key proteins (SPS, UGPase, AGPase and AMY) associated with sugar and starch metabolism during three developmental stages showed that SPS expression level in SC5 was much higher than sugar cassava at three developmental stages, and SPS expression level was highest in SC5 at the expanding period. UGPase expression level in SC5 was higher than sugar cassava in the formation period, but both enzymes had no significant differences in the expanding and mature periods. AGPase expression level in SC5 was much higher than sugar cassava in the formation and the expanding periods, but both enzymes had no significant differences in the mature period.AMY level in SC5 were much higher than that of sugar cassava at three developmental stages.4. SC5 was used as control, comparative proteomics analysis showed that 8 differential proteins spots associated with glycolysis, citric acid cycle and energetic are revealed in sugar cassava at the formation period, in which 4 were up-regulated and 4 were down-regulated.9 differential protein spots associated with glycolysis, citric acid cycle and energetic pathway were found in sugar cassava at the expanding period, in which 6 were up-regulated and 3 were down-regulated. AGPase associated with starch accumulation was down-regulated in sugar cassava at the expanding period.11 differential protein spots associated with glycolysis, citric acid cycle and energetic pathway were detected in sugar cassava at the expanding period, in which 9 were up-regulated and 2 were down-regulated, while SPS associated with starch degration was down-regulated in sugar cassava at the mature period. UGPase associated with sucrose degradation was up-regulated in sugar cassava at the mature period.5. Based on protein-protein interaction database of Arabidopsis, STRING 10 online software was used to build the biological network of protein-protein interaction about differential proteins of sugar cassava and SC5 in the mature period to understand the molecular mechanism of sugar and starch metabolism in sugar cassava. The regulatory network contains 41 protein nodes and 102 kinds of protein-protein interaction relationships.In the network, the proteins with highly interactions were associated with chaperones(10), carbohydrate and energy metabolism (6), detoxifying and antioxidant (3), protein biosynthesis (2), DNA and RNA metabolism (1). The proteins related to carbon and energy metabolism contained enolase 2, triosephosphate isomerase, succinate dehydrogenase flavoprotein subunit, ATP synthase subunit beta vacuolar and pyruvate dehydrogenase. The proteins linked to detoxifying and antioxidant contained monodehydroascorbate reductase family protein, NADP-dependent malic enzyme and malic enzyme. The proteins related to protein biosynthesis contained elongation factor and protein disulfide-isomerase. The proteins associated with DNA and RNA metabolism was nucleoside diphosphate kinase B. All the proteins involved in hexos metabolism were up-regulated, which will improve hexose degradation metabolism and reduced starch accumulation in tuberous root of sugar cassava. |
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