PubMed Journals: J Biol Chem
Source: PMID: 9837919
⇦ ⇨ J Biol Chem. 1998 Dec 11;273(50):33423-8.
Role of asparagine-linked oligosaccharides
in protein folding, membrane targeting,
and thyrotropin and autoantibody binding
of the human thyrotropin receptor.
Nagayama Y(1), Namba H, Yokoyama N, Yamashita
S, Niwa M.
(1) Department of Pharmacology 1,
Nagasaki University School of Medicine,
Nagasaki, 852-8523, Japan.
The amino-terminal ectodomain of thyrotropin (TSH)
receptor (TSHR) is heavily glycosylated with
asparagine-linked (N-linked) oligosaccharides.
The present studies were designed to evaluate
how acquisition and processing of N-linked
oligosaccharides play a role in the functional
maturation of human TSHR. A glycosylation
inhibitor tunicamycin, which inhibits the
first step of N-linked glycosylation (acquisition
of N-linked oligosaccharides), and a series
of mutant Chinese hamster ovary (CHO)-Lec
cells defective in the different steps of
glycosylation processing were used. Inhibition
of acquisition of N-linked oligosaccharides
by tunicamycin treatment in CHO cells stably
expressing TSHR produced nonglycosylated
TSHR, which was totally nonfunctional. In
contrast, all of the TSHRs synthesized in
mutant CHO-Lec1, 2, and 8 cells (mannose-rich,
sialic acid-deficient, and galactose-deficient
oligosaccharides, respectively) bound TSH
and produced cAMP in response to TSH with
an affinity and an EC50 similar to those
in TSHR expressed in parental CHO cells
(CHO-TSHR; sialylated oligosaccharides).
However, Lec1-TSHR and Lec2-TSHR were not
efficiently expressed on the cell surface,
whereas the expression levels of Lec8-TSHR
and CHO-TSHR were essentially identical.
All of the TSHRs expressed in CHO-Lec cells
cleaved into two subunits. Finally, anti-TSHR
autoantibodies from Graves' patients interacted
with all of the TSHRs harboring different
oligosaccharides to a similar extent. These
data demonstrate that acquisition and processing
of N-linked oligosaccharides of TSHR appear
to be essential for correct folding in the
endoplasmic reticulum and for cell surface
targeting in the Golgi apparatus. We also
show that complex type carbohydrates are
not crucially involved in the interaction
of TSHR with TSH and anti-TSHR autoantibodies.
PMID: 9837919 [Indexed for MEDLINE]