Functional analysis of the promoter of the MdFRK2 gene encoding a high-affinity fructokinase in apple (malus × domestica)

Our previous studies have revealed that the MdFRK2 gene is a key gene in determining fructokinase (FRK) activity and fructose metabolism. However, the transcriptional regulation mechanism of MdFRK2 is largely unknown. In this study, we cloned the 1730 bp 5′ flanking region of the MdFRK2 gene (MdFRK2P) from 'Gala' apple plants and constructed the pBI121-MdFRK2P expression vector by fusing the putative promoter with the GUS reporter gene and stably transforming it into Arabidopsis thaliana. Subsequent GUS staining indicated that MdFRK2 promoter expression was extremely high in active sink tissues (shoot tips). Bioinformatics analysis showed a large number of cis-acting elements associated with plant hormone regulation, light responsiveness and abiotic stress located within the 1730 bp 5′ regulatory region. To identify the critical regulatory elements that control the transcription of the MdFRK2 gene, four different lengths of 5′ deletions (-1730, -1500, -1200 and -600 bp) were fused to the GUS gene and transformed into tobacco plants. The -600 bp deletion had the highest activity. Furthermore, the MdFRK2 promoter region was strongly induced by exogenous fructose, exogenous glucose and drought stress. These findings provide essential information for studying the transcriptional regulation mechanism of MdFRK2.