Genome-Wide Identification And Functional Analysis Of Sugar Transporter Genes In Cassava(Manihot Esculenta)

Sugar transporters,which mediate the transport of sugar and participate in the transport and distribution of carbohydrates between source and sink tissues,play an important role in plant growth and development.As a kind of root tuber crops,cassava's yield is mainy related to the number of root tubers and the starch content.The study of cassava sugar transporter genes is expected to provide new insights for understanding the sugar unloading pathway and the mechanism of root tubers formation and the high efficient starch synthesis.In this paper,the sugar transporter genes were indentified,further more,their cluster evolution,gene structure,protein conserved domains,gene duplication model,natural selection pressure were analyzed with the use of cassava genome data.In addition,the MeSTPs and MeSUTs expression patterns in different tissues and developmental stages of cassava were analyzed by using real-time PCR.The subcellular localization analysis and functional verification of 16 cassava STPs were carried out by means of the transient expression system of cassava protoplasts and functional complementation of yeast mutants.The main results are as follows:(1)A total of 69 open reading frames(ORFs)encoding putative sugar transporter proteins were identified in the cassava.Phylogenetic analysis of the 69 identified nucleotide sequences reveals that sugar transporters could be classified into eight separate subfamilies.There are twenty members in the subfamily of STP,fifteen members in the subfamily of SFP,ten members in subfamily of PLT,six members in the subfamily of TMT and pGlcT,five members in the subfamily of SUT and INT,and two members in the subfamily of VGT.The members of each subfamily are conservative in gene structure and protein domain,but with great differences among the subfamilies.Gene duplication event analysis indicated that whole genome duplication(WGD)and segmental duplication play key roles in sugar transporter gene amplification,followed by tandem duplication.Estimation of positive selection at codon sites of sugar transporter paralog pairs indicated that all duplicated gene pairs have evolved under purify selection.(2)The expression of sugar transporter gene in different tissues and developmental stages of cassava shows great temporal and spatial specificity:In the cassava shoots apical meristm,MeSTP2 and MeSUT4 was the highest;In leaves,the expression of MeSTP17 and MeSUT5 was the highest;In bud,MeSTP2 and MeSUT4 expression was the highest;In the stem,the highest expression genes were MeSTP17 and MeSUT5;In the fiberous roots,MeSTP19 and MeSUT5 expression was the highest;In root tubers,the expression of MeSTP2 and MeSUT3 was the highest;In the apical meristem,MeSTP7 and MeSUT1 have high abundance expression;The analysis of the expression of STPs and SUTs in the different developmental stages of cassava roots showed that the gene expression was highly temporal specific patterns.All genes were highly expressed in early tuberous roots,and the genes expression decreased with the development and maturation of tuberous roots.(3)Sixteen MeSTPs were amplified from cassava and then transferred into yeast mutant EBY.VW 4000 to study the functions of the sugar transporter.The results.show that MeSTP2 and MeSTP19 can transport mannose,galactose,xylose,glucose and fructose with high capacity;MeSTP3,MeSTP5 and MeSTP16 showed galactose specific transport,but their transport capacity was weak;MeSTP7,MeSTP12 and MeSTP 17 have strong ability to transport mannose,galactose,glucose and fructose,except for xylose;MeSTP4,MeSTP10,MeSTP13,MeSTP14 and MeSTP20 showed no ability to transport any monosaccharide.In addition,the proteins subcellular localization in cassava protoplasts determined that all MeSTP were cell membrane localized.In summary,cassava owns a large number of sugar transporter genes that are involved in sugar transport in various tissues.The expression patterns of MeSTP and MeSUT genes during cassava root development suggested that the sugar unloading pathway of cassava had undergone a transformming from apoplastic pathway to symplast pathway.All MeSTPs are localized on the cell membrane and most members exhibit a broad spectrum of transport characteristics for monosaccharide substrates.MeSTPs are responsible for transport of apoplastic monosaccharides into the membrane,which may be involved in the information of cassava tubers.This study is important to elucidate the biological functions of sugar transporters in the development of cassava tubers and to provide theoretical basis and candidate gene resources for the improvement of cassava varieties.