Identification and pharmacological correction of a membrane trafficking defect associated with a mutation in the sulfonylurea receptor causing familial hyperinsulinism
Cj. Partridge et al., Identification and pharmacological correction of a membrane trafficking defect associated with a mutation in the sulfonylurea receptor causing familial hyperinsulinism, J BIOL CHEM, 276(38), 2001, pp. 35947-35952
Persistent hyperinsulinemic hypoglycemia of infancy (PHHI) is a genetic dis
order characterized by excess secretion of insulin and hypoglycemia. In mos
t patients, the disease is caused by mutations in sulfonylurea receptor-1 (
SUR1), which, in association with Kir6.2, constitutes the functional ATP-se
nsitive potassium (K-ATP) channel of the pancreatic beta -cell. Previous st
udies reported that coexpression of the PHHI mutant R1394H-SUR1 with Kir6.2
in COS cells produces no functional channels. To investigate if the loss o
f function could be due to impaired trafficking of mutant channels to the c
ell membrane, we have cotransfected wild-type and mutant SUR1 subunits with
Kir6.2 into HEK293 cells and examined their cellular localization by immun
ofluorescent staining. Our results show that unlike the wildtype subunits,
which showed fluorescence at the cell surface, the mutant subunits displaye
d fluorescence in punctate structures. Co-immunostaining with antibodies ag
ainst organelle-specific marker proteins identified these structures as the
trans-Golgi network. Limited localization in clathrin-positive, but transf
errin receptor-negative vesicles was also observed. The post-endoplasmic re
ticulum localization suggests that the mutation does not impair the folding
and assembly of the channels so as to cause its retention by the endoplasm
ic reticulum. Diazoxide, a KATP channel opener drug that is used in the tre
atment of PHHI, restored the surface expression in a manner that could be p
revented by the channel blocker glibenclamide. When expressed in Xenopus oo
cytes, R1394H-SUR1 formed functional channels with Kir6.2, indicating that
the primary consequence of the mutation is impairment of trafficking rather
than function. Thus, our data uncover a novel mechanism underlying the the
rapeutic action of diazoxide in the treatment of PHHI, i.e. its ability to
recruit channels to the membrane. Furthermore, this is the first report to
describe a trafficking disorder effecting retention of mutant proteins in t
he trans-Golgi network.