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Human leukemia cell lines bind basic fibroblast growth factor (FGF) on FGF
receptors and heparan sulfates: downmodulation of FGF receptors by phorbol
ester
JP Liuzzo and D Moscatelli
Department of Cell Biology, New York University Medical Center, New York
10016, USA.
Basic fibroblast growth factor (bFGF) has been identified as an important
cytokine for blood cells. To determine whether hematopoietic cells have
receptors that recognize bFGF, the ability of human leukemia cell lines to
bind 125I-bFGF was investigated. Specific bFGF-binding sites were
identified on K562 and HL60 cells, but not on U937 cells. DAMI cells bound
low amounts of 125I-bFGF specifically. Binding of 125I- bFGF to K562 cell
surfaces was reduced in a dose-dependent manner by unlabeled bFGF or by
heparin. Scatchard analysis of binding to K562 cells revealed two classes
of binding sites: 1,650 high affinity binding sites per cell with a
dissociation constant (kd) of 192 pmol/L, and 36,600 low affinity sites per
cell with a kd of 9.3 nmol/L. Chemical crosslinking experiments with K562,
HL60, and DAMI cells revealed receptor-growth factor complexes with
molecular masses of 140 to 160 kD, similar in size to complexes formed by
known receptor species. Binding of 125I-bFGF to K562 cells was sensitive to
heparinase treatment but not to chondroitinase treatment, suggesting that
heparan sulfate proteoglycans (HSPGs) may be responsible for the low
affinity binding sites. To further investigate whether K562 cells make
HSPG, the incorporation of 35SO4 into proteoglycans was assessed.
Metabolically labeled cell-surface proteoglycans with molecular masses of
180 to 300 kD were identified in K562 cells. These proteoglycans were
sensitive to heparinase, demonstrating that K562 cells synthesize
bFGF-binding HSPG. Treatment of K562 cells with
phorbol-12-myristate-13-acetate (PMA) caused a loss of bFGF-binding
capacity. This decreased binding capacity reflected a rapid loss of high
affinity receptors. The ability to form bFGF-receptor complexes decreased
by 65% to 70% within 1 hour and declined continuously thereafter. The
decrease in binding of bFGF was not due to an autocrine downregulation of
bFGF receptors, because there was no increase in bFGF after PMA treatment
as detected by Western blotting, and suramin, which blocks bFGF binding to
receptors, did not prevent the loss of receptors after exposure to PMA. In
addition, inhibitors of either protein synthesis or protease activity did
not prevent the loss of bFGF receptors in PMA-treated cells. In summary,
this work demonstrates that leukemia cell lines have receptors that
specifically bind bFGF and supports the hypothesis that bFGF acts directly
on certain blood cells to stimulate their proliferation.
Volume 87,
Issue 1,
pp. 245-255,
01/01/1996
Copyright © 1996 by The American Society of Hematology
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