title: Cloning, characterization, and expression of glucose transporter 2 in the freeze-tolerant wood frog, Rana sylvatica.
abstract: BACKGROUND
  The essential role of glucose transporter 2 (GLUT2) in glucose homeostasis has been extensively studied in mammals; however, little is known about this important protein in lower vertebrates. The freeze-tolerant wood frog (Rana sylvatica), which copiously mobilizes glucose in response to freezing, represents an excellent system for the study of glucose transport in amphibians.
  
  
  METHODS
  GLUT2 was sequenced from northern and southern phenotypes of R. sylvatica, as well as the freeze-intolerant Rana pipiens. These proteins were expressed and functionally characterized in Xenopus oocytes. Abundance of GLUT2 in tissues was analyzed using immunoblotting techniques.
  
  
  RESULTS
  GLUT2s cloned from these anurans encoded proteins with high sequence homologies to known vertebrate GLUT2s and had similar transport properties, although, notably, transport of the glucose analog 3-O-methyl-d-glucose (3-OMG) was strongly inhibited by 150mM urea. Proteins from all study subjects had similar affinity constants (~12mM) and other kinetic properties; however, GLUT2 abundance in liver was 3.5-fold greater in northern R. sylvatica than in the southern conspecific and R. pipiens.
  
  
  CONCLUSIONS
  Our results indicate that amphibian GLUT2s are structurally and functionally similar to their homologs in other vertebrates, attesting to the conserved nature of this transport protein. The greater abundance of this protein in the northern phenotype of R. sylvatica suggests that these transporters contribute importantly to freezing survival.
  
  
  CONCLUSIONS
  This study provides the first functional characterization of any GLUT isoform from an anuran amphibian and novel insights into the role of these proteins in glucose homeostasis and cryoprotectant mobilization in freeze-tolerant vertebrates.
authors.affiliation: Department of Zoology, Miami University, 700 E. High St., Oxford, OH 45056, USA. Electronic address: rosendaj@miamioh.edu.
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Rosendale
authors.affiliation: Department of Biology, Rivier University, 420 S. Main St., Nashua, NH 03060, USA. Electronic address: bphilip@rivier.edu.
authors.forename: Benjamin N
authors.initials: BN
authors.lastname: Philip
authors.affiliation: Department of Zoology, Miami University, 700 E. High St., Oxford, OH 45056, USA. Electronic address: leere@miamioh.edu.
authors.forename: Richard E
authors.initials: RE
authors.lastname: Lee
authors.affiliation: Department of Zoology, Miami University, 700 E. High St., Oxford, OH 45056, USA. Electronic address: costanjp@miamioh.edu.
authors.forename: Jon P
authors.initials: JP
authors.lastname: Costanzo
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D005615:Freezing; D051273:Glucose Transporter Type 2 / Q000235:genetics*; D008297:Male; D008969:Molecular Sequence Data; D010802:Phylogeny; D011898:Ranidae; D011994:Recombinant Proteins / Q000096:biosynthesis*; D014018:Tissue Distribution; D014508:Urea / Q000494:pharmacology
chemical_list: D051273:Glucose Transporter Type 2; D011994:Recombinant Proteins; D014508:Urea
keywords: Anuran; Cryoprotectant; Ecogeography freeze tolerance; Glucose transport