Isolation And Initial Function Characterization Of The Monosaccharide Transporter Gene MeSTP2 From Manihot Esculenta Crantz

Sugars(sucrose,monosaccharides,polyols,etc.)plays an important role in higher plant growth,development,maturation process.Among the sugars,monosaccharides are mainly transported by monosaccharide transporters(MSTs),which are consist of several subfamilies.While in the subfamilies,sugar transport proteins(STPs)gene subfamily may play an important role in the process of transporting the monosaccharides outside the cell membrane into cells.These monosaccharides can not only served as a substrate in the synthesis of various compounds such as sucrose.starch and so on,but also be involved in the regulation progress of the expression of other genes as a molecule signal worked with a series of hexokinases.In this paper,the MeSTP2 gene was cloned by RT-PCR,and then its function and expression analysis were studied by yeast heterologous expression,subcellular localization,quantitative real-time PCR technology.The main results are as follows:1.MeSTP2 with sequence length of 1 575 bp encodes a protein of 524 amino acids.It belongs to the major facilitator superfamily(MFS)by the conserved domain analysis of its protein sequence,the presence of Sugar_tr domains also suggests it may have sugar transport capability.The hydrophobic prediction showed that the protein had 12 transmembrane structures.Compared monosaccharide transporter proteins with the species of castor,grape and arabidopsis,it found that MeSTP2 was highly homologous with the monosaccharide transporter proteins located on the cell membrane,while,with a homology of 92%,MeSTP2 was most homologous with RCSTC,a monosaccharide transporter protein in castor.2.The heterologous expression analysis suggested that MeSTP2 had a high affinity with D-glucose,D-mannose,D-fructose,D-galactose except D-xylose.3.The vector combined of MeSTP2 and green fluorescent gene was transformed into the yeast cells and cassava protoplasts.The fusion protein was clearly targeted on the cell membrane when observed under the confocal microscope.4.The expression of MeSTP2 in different development periods roots of high and low starch content cassava(KU50.CAS36)were measured.In the two kinds of cassavas,the MeSTP2 expression level declined from the seedling stage to the root formation stage(45 d N100 d);In the root expansion stage(100 d～180 d),the MeSTP2 expression level was increased in CAS36.With the maturation of root,the expression of MeSTP2 became stable;However,regardless of any stage,expression level in phloem was higher than in xylem.Comparing the gene expression in different tissues in the root expansion stage of cassava,MeSTP2 expression was high mainly in the tissues with vigorous metabolism(fibrous root,stem,young leaves ect.).In summary,MeSTP2 is a plasma membrane hexose transporter.It transports several kinds of monosaccharide.MeSTP2 gene is involved in the process of sugar unloading and its expression has relationship with the development of tissues and the sugar metabolisms.