The Journal of Cell Biology, Vol 93, 860-865, Copyright © 1982 by The Rockefeller University Press
Internalization and degradation of cholera toxin by cultured cells: relationship to toxin action
PH Fishman
Using anticholeragen antibodies and 125I-protein A, we developed a specific
and quantitative assay for measuring choleragen on the surfaces of cultured
cells. When neuroblastoma cells containing bound toxin were incubated at 37
degrees C, surface toxin disappeared with a half-life of approximately 2 h
and a significant loss was detected by 10 min. When cells were incubated
with 125I-choleragen in order to measure toxin degradation, cell-associated
radioactivity disappeared with time and a corresponding amount of
TCA-soluble label appeared in the culture medium with a half-life of 4-6 h.
No degradation was detected until 45 min. Although there was a lag of 15
min before bound choleragen activated adenylate cyclase, the enzyme became
maximally activated between 45 and 60 min. Similar results were obtained
with Friend erythroleukemia cells. Internalization, degradation, and
activation all were blocked when the cells were maintained at 4 degrees C.
At 22 degrees C, internalization and activation occurred, albeit at a
slower rate, whereas degradation was effectively inhibited. These results
indicated that choleragen does not have to be degraded by intact cells in
order for it to activate adenylate cyclase. Some internalization of the
toxin, however, appears to precede the activation process.
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