D. Fukushima et al., LOCALIZATION OF TRANSFORMING GROWTH-FACTOR-BETA BINDING-SITE IN BETAGLYCAN - COMPARISON WITH SMALL EXTRACELLULAR-MATRIX PROTEOGLYCANS, The Journal of biological chemistry, 268(30), 1993, pp. 22710-22715
The most abundant binding molecule for transforming growth factor beta
(TGF-beta) on many cell types is betaglycan, a transmembrane proteogl
ycan. To localize the binding site for TGF-beta in betaglycan, parts o
f the betaglycan extracellular portion were expressed as recombinant f
usion proteins in bacteria and tested for their ability to compete for
the binding of TGF-beta1 to Hep G2 cells. One fragment encompassing 2
26 residues near the transmembrane domain (amino acids 543-769) (Lopez
-Casillas, F., Cheifetz, S., Doody, J., Andres, J. L., Lane, W. S., an
d Massague, J. (1991) Cell 67, 785-795) was active, whereas fusion pro
teins representing the other parts of the betaglycan ectodomain were i
nactive. Affinity measurements revealed two classes of binding sites (
K(d) = 3.9 nM and K(d) = 145 nm) for the active fusion protein. The bi
nding of the betaglycan fusion protein to immobilized TGF-beta-1 was i
nhibited by fusion proteins representing the core proteins of the smal
l interstitial proteoglycans decorin, biglycan and fibromodulin, each
also known to bind TGF-beta. The effective concentrations of TGF-beta
for binding to these other proteoglycans were similar to those require
d for binding to betaglycan, indicating similar affinities for the bin
ding of proteoglycans and betaglycan. Affinity cross-linking showed th
at, at low concentrations, the betaglycan fragment enhanced the bindin
g of TGF-beta to the type II receptor and to endogenous betaglycan but
had no effect on the binding to type I receptor. At high concentratio
ns, the TGF-beta binding fragment inhibited the binding of TGF-beta to
all these receptors. The fragment enhanced the activity of TGF-beta i
n mink lung cell bioassay at all active concentrations. The results in
dicate that betaglycan and the decorin type proteoglycans all bind to
the same or closely spaced sites in TGF-beta and compete with one anot
her for the binding. In addition, betaglycan may cooperate with the ty
pe II receptor on TGF-beta binding.