A hexokinase gene named MdHXK1(MDP0000309677) was cloned from ‘Gala' apple(Malus × domestica Borkh.). Sequence analysis showed that the MdHXK1 gene was 1 497 bp long and encoded 499 amino acids. The predicte...A hexokinase gene named MdHXK1(MDP0000309677) was cloned from ‘Gala' apple(Malus × domestica Borkh.). Sequence analysis showed that the MdHXK1 gene was 1 497 bp long and encoded 499 amino acids. The predicted molecular mass of this protein was 54.05 kD, and the pI was 5.76. A phylogenetic tree indicated apple MdHXK1 exhibited the highest sequence similarity to Pyrus bretschneideri PbHXK1. Analysis of the functional domain showed that the MdHXK1 protein included two conserved kinase domains. The prediction of subcellular localization suggested that the MdHXK1 protein was mainly localized in the cytoplasm. There was an indication that MdHXK1 existed as one copy in the apple genome by Southern blotting. Silico analysis suggested that the promoter sequence contained several typical cis-acting elements, including defense, sugar signaling and phytohormone responsive elements. Quantitative real-time PCR analysis demonstrated that the MdHXK1 gene was mainly expressed in stem and flower tissues. During the development of apple fruits, the expression of the MdHXK1 gene initially increased and then decreased. The changes on Glc phosphorylation relative activity and glucose concentration showed the same trend. In addition, the expression of this gene was induced by salt stress, low temperature, and abscisic acid(ABA). Finally, we obtained and purified the fused MdHXK1 protein by recombinant prokaryotic expression. Studies have demonstrated that MdHXK1 may participate in sugar metabolism in apple fruits. Enzyme encoded by MdHXK1 is a key factor in the mediation of sugar accumulation. Recently,researchers on hexokinase at home and abroad mainly focused on model plants, such as Arabidopsis, tobacco and rice, but orchard fruit like apple were underresearched. Our research established the foundation for the further study of the functions of MdHXK1.展开更多
Calcium(Ca^(2+)), a crucial second messenger in plants, is involved in diverse signaling pathways including biotic and abiotic stress responses. However, the functions of most calcium sensors including calcium-depende...Calcium(Ca^(2+)), a crucial second messenger in plants, is involved in diverse signaling pathways including biotic and abiotic stress responses. However, the functions of most calcium sensors including calcium-dependent protein kinases(CDPKs), calmodulins(Ca Ms), calmodulinlike proteins(CMLs), and calcineurin B-like proteins(CBLs) remain limited in plants, particularly in woody plants. Currently, a total of 83 CMLs and seven Ca Ms were discovered in the apple genome in this study. Functional domain analysis showed that these CMLs and Ca Ms contain a highly conserved EF-hand motif. q RT-PCR assays indicated that these CMLs were expressed ubiquitously in apple, including in the roots, stems,leaves, flowers, and fruits, and also possessed spatial specificity. Finally, most of these CMLs were induced by hormonal and abiotic stress, suggesting their potential roles in the regulation of growth, development, and the stress response in apple. In summary, our findings provide potential evidence that apple CMLs may be involved in abiotic stress and the regulation of plant growth and development.展开更多
基金supported by the National Science Foundation for Distinguished Young Scholars of China(Grant No.31325024)Innovation Team Project of Shandong Modern Agricultural Industry Technology System(SDAIT-03-022-03)
文摘A hexokinase gene named MdHXK1(MDP0000309677) was cloned from ‘Gala' apple(Malus × domestica Borkh.). Sequence analysis showed that the MdHXK1 gene was 1 497 bp long and encoded 499 amino acids. The predicted molecular mass of this protein was 54.05 kD, and the pI was 5.76. A phylogenetic tree indicated apple MdHXK1 exhibited the highest sequence similarity to Pyrus bretschneideri PbHXK1. Analysis of the functional domain showed that the MdHXK1 protein included two conserved kinase domains. The prediction of subcellular localization suggested that the MdHXK1 protein was mainly localized in the cytoplasm. There was an indication that MdHXK1 existed as one copy in the apple genome by Southern blotting. Silico analysis suggested that the promoter sequence contained several typical cis-acting elements, including defense, sugar signaling and phytohormone responsive elements. Quantitative real-time PCR analysis demonstrated that the MdHXK1 gene was mainly expressed in stem and flower tissues. During the development of apple fruits, the expression of the MdHXK1 gene initially increased and then decreased. The changes on Glc phosphorylation relative activity and glucose concentration showed the same trend. In addition, the expression of this gene was induced by salt stress, low temperature, and abscisic acid(ABA). Finally, we obtained and purified the fused MdHXK1 protein by recombinant prokaryotic expression. Studies have demonstrated that MdHXK1 may participate in sugar metabolism in apple fruits. Enzyme encoded by MdHXK1 is a key factor in the mediation of sugar accumulation. Recently,researchers on hexokinase at home and abroad mainly focused on model plants, such as Arabidopsis, tobacco and rice, but orchard fruit like apple were underresearched. Our research established the foundation for the further study of the functions of MdHXK1.
基金grants from NSFC(31601728,31272142)Shandong Province(ZR2016CQ13,SDAIT-06-03)+1 种基金Young Scientists Funds of Shandong Agricultural University(564024)Youth Science and Technology Innovation Fund of Shandong Agricultural University(24024)
文摘Calcium(Ca^(2+)), a crucial second messenger in plants, is involved in diverse signaling pathways including biotic and abiotic stress responses. However, the functions of most calcium sensors including calcium-dependent protein kinases(CDPKs), calmodulins(Ca Ms), calmodulinlike proteins(CMLs), and calcineurin B-like proteins(CBLs) remain limited in plants, particularly in woody plants. Currently, a total of 83 CMLs and seven Ca Ms were discovered in the apple genome in this study. Functional domain analysis showed that these CMLs and Ca Ms contain a highly conserved EF-hand motif. q RT-PCR assays indicated that these CMLs were expressed ubiquitously in apple, including in the roots, stems,leaves, flowers, and fruits, and also possessed spatial specificity. Finally, most of these CMLs were induced by hormonal and abiotic stress, suggesting their potential roles in the regulation of growth, development, and the stress response in apple. In summary, our findings provide potential evidence that apple CMLs may be involved in abiotic stress and the regulation of plant growth and development.
