Premium
Dynamic modulation of the cell surface expression of the granulocyte‐macrophage colony‐stimulating factor receptor
Author(s) -
Khwaja Asim,
Carver Julia,
Jones H. Mark,
Linch David C.
Publication year - 1993
Publication title -
british journal of haematology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.907
H-Index - 186
eISSN - 1365-2141
pISSN - 0007-1048
DOI - 10.1111/j.1365-2141.1993.tb08643.x
Subject(s) - receptor , ligand (biochemistry) , cell surface receptor , microbiology and biotechnology , cell , granulocyte macrophage colony stimulating factor , binding site , biology , chemistry , biophysics , biochemistry , cytokine , immunology
Summary. The GM‐CSF receptor (GM‐CSFR) is composed of α and β subunits. Surface expression of the α chain alone leads to low affinity GM‐CSF binding and of both subunits to high affinity binding: the β chain is required for transducing a proliferative signal. Studies of GM‐CSFR expression have concentrated largely on static events occurring under conditions of binding equilibrium. We have examined the dynamic regulation of high and low affinity GM‐CSFR expression in neutrophils (1100 ± 200 R/cell, K D 50 ± 15 pm) and a GM‐CSF dependent human leukaemic cell line, TF‐l (2000 ± 450 R/cell K D 15 ± 5 PM) and 8600 ± 150 R/cell K D 1.8 ± 0.3 nm). The addition of GM‐CSF to TF‐1 cells (350 PM, 4 h at 37°C) caused a reduction in subsequent binding of 125 I‐GM‐CSF at low ligand concentration (100 pm) (following a low pH wash to remove surface bound ligand) to 16 ± 4% and a reduction in binding at high ligand concentration (2 nm 125 I‐GM‐CSF) to 36 ± 9% of control. Scatchard analysis showed complete down‐regulation of high affinity GM‐CSFR and a significant reduction in low affinity GM‐CSFR. In neutrophils, concentration‐response curves of ligand induced receptor down‐regulation at 37°C showed that observed down‐modulation was more than 10‐fold greater than predicted by static equilibrium binding data and correlated closely with GM‐CSF priming of the neutrophil respiratory burst. The addition of IL‐3 to TF‐1 cells at 37°C reduced 100 PM 125 I‐GM‐CSF binding to 18 ± 4% and 2 nm 125 I‐GM‐CSF binding to 46 ± 5% of control. TF‐1 cells, but not neutrophils, were able to re‐express GM‐CSFR following removal of GM‐CSF from medium. TF‐1 proliferation assays showed that pulsed GM‐CSF (0.35–3.5 nm) for up to 4 h did not cause a significant increase in 3 H‐thymidine incorporation which required the continued presence of GM‐CSF (control 2875 ± 208 cpm, pulsed GM‐CSF 5 ng/ml 49 72 ± 1344. continuous GM‐CSF 5 ng/ml 17249 ± 2982). Therefore, proliferation of TF‐1 cells required the continued presence of GM‐CSF at a time when there was no detectable surface high affinity GM‐CSFR. This shows that signal transduction can take place via the GM‐CSFR at a time when there is no detectable high affinity GM‐CSF binding and introduces a further layer of complexity in the analysis of GM‐CSF receptor function.