Characterization of two novel forms of the rat sulphonylurea receptor SUR1A2 and SUR1BDelta31.
1. The ATP-sensitive potassium channel (K(ATP)) of pancreatic beta-cells is composed of the sulphonylurea-binding protein, SUR1, and the inwardly rectifying K(+) channel subunit, Kir6.2. We have characterized two novel isoforms of rat SUR1 in the RINm5F insulin-secreting cell line. 2. SUR1A2 is an allelic variant with a single amino acid change in the first nucleotide-binding domain. Coinjection of SUR1A2 plus Kir6.2 into Xenopus oocytes or expression of a SUR1A2-Kir6.2 tandem in HEK-293 cells yielded large currents with characteristics similar to the wild-type K(ATP) channel. 3. SUR1BDelta31, detected in several human tissues, is a splice variant of the rat SUR1 gene that lacks exon 31 of the corresponding human SUR1 gene. SUR1BDelta31 lacks the TM16-TM17 transmembrane-spanning helices leading to a protein with a different transmembrane topology. Coinjection of SUR1BDelta31 plus Kir6.2 into Xenopus oocytes or expression of the Kir6.2/SUR1BDelta31 tandem construct in HEK-293 cells did not result in any current, and a surface expression assay indicated that this channel does not reach the plasma membrane. 4. SUR1A2 and SUR1A1 proteins expressed in HEK-293 cells display similar binding affinities for [(3)H]-glibenclamide, while SUR1BDelta31 shows a 500-fold lower affinity. 5. These findings confirm that TM16-TM17 of SUR1 are important for high-affinity glibenclamide binding and that their deletion impairs trafficking of the K(ATP) channel to the surface membrane.
Gros
Laurent
L
Institut National de la Santé et de la Recherche Médicale INSERM U376, CHU Arnaud-de-Villeneuve, 34295 Montpellier Cedex, France. gros@igm.cnrs-mop.fr
Trapp
Stefan
S
Dabrowski
Michael
M
Ashcroft
Frances M
FM
Bataille
Dominique
D
Blache
Philippe
P
D018528:ATP-Binding Cassette Transporters; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D005905:Glyburide / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D009865:Oocytes; D009928:Organ Specificity; D018408:Patch-Clamp Techniques; D016133:Polymerase Chain Reaction; D015221:Potassium Channels / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D024661:Potassium Channels, Inwardly Rectifying / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D020033:Protein Isoforms; D011869:Radioligand Assay; D051381:Rats; D011955:Receptors, Drug / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D064233:Sulfonylurea Receptors; D014981:Xenopus
C577507:ABCC8 protein, human; D018528:ATP-Binding Cassette Transporters; C577509:Abcc8 protein, rat; D007328:Insulin; D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D020033:Protein Isoforms; D011955:Receptors, Drug; D064233:Sulfonylurea Receptors; D000255:Adenosine Triphosphate; D005905:Glyburide