The Journal of Cell Biology, Vol 95, 697-703, Copyright © 1982 by The Rockefeller University Press
125I-thrombin binds to clustered receptors on noncoated regions of mouse embryo cell surfaces
DH Carney and JS Bergmann
We used electron microscope autoradiography (EMAR) to visualize the
interaction of 125I-thrombin with its surface receptors on mouse embryo
(ME) cells. Autoradiographic grains were spaced over the surface of cells
in a periodic nonrandom pattern, indicating 125I-thrombin association with
clusters of thrombin receptors. The grain spacing varied slightly from cell
to cell, indicating subpopulations of cells with different numbers of
thrombin receptors. The average distance between grains on ME cells after
binding 125I-thrombin (125 ng/ml) at 37 degrees C was 1.65 +/- 0.49
microns. The average distance between grains on prefixed cells and cells
incubated with 125I-thrombin at 4 degrees C was not significantly different
from that observed at 37 degrees C. This indicates that thrombin receptors
are clustered before thrombin binding and that the thrombin receptor
aggregates do not redistribute into large aggregates on the surface of
cells subsequent to thrombin binding. The number of grains per cluster also
does not change under these three binding conditions. Thus, the number of
occupied receptors in each cluster appears to be constant. On the basis of
the average grain number and spacing, we estimate that each cluster is
approximately 400 nm in diameter containing approximately 550
thrombin-binding sites. These receptor-clusters are not associated with
specialized structures or coated regions of the membrane. Additionally,
grains observed within cells were not found associated with coated
vesicles. Therefore, neither the clustering patterns nor internalization of
125I-thrombin are characteristic of molecules which bind to receptors and
are internalized by receptor-mediated endocytosis.
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