Functional expression and characterisation of a gut facilitative glucose transporter, NlHT1, from the phloem-feeding insect Nilaparvata lugens (rice brown planthopper).
Phloem-sap feeding Hemipteran insects have access to a sucrose-rich diet but are dependent on sucrose hydrolysis and hexose transport for carbon nutrition. A cDNA library from Nilaparvata lugens (rice brown planthopper) was screened for clones encoding potential transmembrane transporters. A selected cDNA, NlHT1, encodes a 53kDa polypeptide with sequence similarity to facilitative hexose transporters of eukaryotes and prokaryotes, including GLUT1, the human erythrocyte hexose transporter. NlHT1 was expressed as a recombinant protein in the methylotropic yeast Pichia pastoris, and was identified in a membrane fraction isolated from transformed yeast cells. Transport experiments using membrane vesicles containing NlHT1 showed that the protein is a saturable, sodium independent transporter, with a relatively low affinity for glucose (K(m) 3.0mM), which can be inhibited by cytochalasin B. Competition experiments with fructose demonstrate NlHT1 is glucose specific. In situ localisation studies revealed that NlHT1 mRNA is expressed in N. lugens gut tissue, mainly in midgut regions, and that expression is absent in hindgut and Malpighian tubules. NlHT1 is therefore likely to play an important role in glucose transport from the gut, and in carbon nutrition in vivo. This is the first report of a facilitative glucose transporter from a phloem-feeding insect pest.
Price
Daniel R G
DR
School of Biological and Biomedical Sciences, University of Durham, Durham DH1 3LE, UK. d.r.g.price@durham.ac.uk
Wilkinson
Hillary S
HS
Gatehouse
John A
JA
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D005247:Feeding Behavior; D051246:Glucose Transport Proteins, Facilitative / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D006430:Hemiptera / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D019476:Insect Proteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D052585:Phloem / Q000378:metabolism; D010843:Pichia / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
D051246:Glucose Transport Proteins, Facilitative; D019476:Insect Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins