Abstract
To investigate the functions of fructokinase (FRK) in apple (Malus domestica) carbohydrate metabolism, we cloned the coding sequences of MdFRK1 and MdFRK2 from the ‘Royal Gala’ apple. The results showed that MdFRK2 expression was extremely high in shoot tips and young fruit. Analyses of heterologously expressed proteins revealed that MdFRK2 had a higher affinity for fructose than did MdFRK1, with Km values of 0.1 and 0.62 mM for MdFRK2 and MdFRK1, respectively. The two proteins, however, exhibited similar Vmax values when their activities were significantly inhibited by high concentrations of fructose. MdFRK2 ectopic expression was associated with a general decrease in fructose concentration in transgenic lines. In leaves, increased FRK activity similarly resulted in reduced concentrations of glucose and sucrose but no alterations in sorbitol concentration. When compared with those in the untransformed control, genes involved in sorbitol synthesis (A6PR) and the degradation pathway (SDH1/2) were significantly upregulated in transgenic lines, whereas those involved in sucrose synthesis (SPS1) and other degradation processes (SUSY4, NINV1/2, and HxK2) were downregulated. The activity of enzymes participating in carbohydrate metabolism was proportional to the level of gene expression. However, the growth performance and photosynthetic efficiency did not differ between the transgenic and wild-type plants. These results provide new genetic evidence to support the view that FRK plays roles in regulating sugar and sorbitol metabolism in Rosaceae plants.
Highlights
Active leaves produce various forms of carbohydrates, which comprise primarily sucrose and starch in most plant species
Our previous studies indicated that MdFRK2 might be a key gene in determining FRK activity and fructose homeostasis[30,31,32]
To investigate the function of the MdFRK2 gene, we cloned the coding sequence of MdFRK2 from ‘Royal Gala’ apple plants, and the MdFRK1 coding sequence was cloned as a reference gene
Summary
Active leaves produce various forms of carbohydrates, which comprise primarily sucrose and starch in most plant species. These compounds serve as energy and carbon sources for metabolism, including the understanding the mechanisms of carbohydrate metabolism is important for exploiting sugar metabolism to improve plant growth and crop quality. In the mature green leaves of most plants, sucrose is the main end product of photosynthesis in mesophyll cells[4]. 3.2.1.26), including cell wall invertase (CWINV), neutral invertase (NINV), and vacuolar invertase (VINV) or to fructose and UDP-glucose (UDPG) by sucrose synthase (SUSY; EC 2.4.1.13)[4]. HxK and FRK are central in sugar metabolism and homeostasis
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