The Journal of Cell Biology, Vol 94, 12-19, Copyright © 1982 by The Rockefeller University Press
Temperature shifts induce the selective loss of alveolar-macrophage plasma membrane components
J Kaplan and EA Keogh
A shift in the incubation temperature of rabbit alveolar macrophages (0
degree C leads to 37 degrees C leads to 0 degree C) resulted in a 40- 60%
reduction in the ability of cells to bind alphamacroglobulin. 125I- trypsin
complexes (alphaM. 125I-T). The reduction in binding activity did not
reflect a disruption of cell integrity since the levels of intracellular
components (lactate dehydrogenase, beta-N-acetyl- hexosaminidase) or other
plasma membrane components (alkaline phosphodiesterase) were unaltered.
Analysis of receptor-ligand interaction indicated that the temperature
shift effected a decline in receptor number rather than an alteration in
ligand-receptor affinity. Studies indicated that a temperature shift
resulted in the loss of unoccupied receptors, and that ligand bound to
receptors was not lost. However, after ligand internalization, receptors
were removed by the temperature shift. The rate of receptor loss was
maximal when cells were incubated at temperatures greater than 24 degrees
C. Receptor loss was not prevented by treatment of cells with colchicine,
cytochalasin B, or N-ethylamaleimide, but was prevented by treatment with
the cross- linking agent paraformaldehyde. Data indicate that the reduction
in alphaM. 125I-T binding activity resulted from shedding of receptors into
the media since media obtained from temperature-shifted cells contained
material that competed with cell-bound receptors for alphaM. 125I-T.
Additionally, binding of alphaM. 125I-T was diminished on membrane
fragments obtained from temperature-shifted cells. Incubation with Triton
X-100, of cells whose receptors were occupied with alphaM. 125I-T, led to
the extraction of 40% of cell-bound activity. However, no radioactivity was
extracted from cells labeled with alphaM. 125I-T after a temperature shift.
Measurement of ligand accumulation by control and temperature-shifted cells
incubated at 20 degrees C indicated that control cells exhibited a
subpopulation of receptors capable of binding ligand but only slowly
internalizing it. This subpopulation was not present on temperature-shifted
cells. These results indicate that surface receptors for alphamacroglobulin
. protease complexes are heterogeneous and that the temperature shift
resulted in the selective loss of membrane components.
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