To determine if a portion of the Tf-FITC and LDLR pools were indeed
trapped in peripheral SEs with high (10 lM) ONO, HeLa cells
were pre-treated with or without 10 lM ONO for 10 min, then
pulse-labeled with Tf-FITC for 45 min at 37 C, and subjected to
various chase protocols. In cells pre-treated with 10 lM ONO for
10 min followed by continuous uptake of Tf-FITC for 45 min, Tf
and LDLR were located in puncta throughout the cytoplasm corresponding
to SEs (Fig. 2A and B). Additional evidence for this conclusion
was obtained by showing that both Tf-FITC and LDLR
redistributed to the central ERC when cells were washed free of
ONO and then chased for just 20 min (Fig. 2C and D). However, if
cells were kept in 10 lM ONO during the chase, then Tf and LDLR
remained present in the more peripheral SEs (Fig. 2E and F). These
results show that 10 lM ONO causes a reversible block in the export
of both Tf and LDLR out of peripherally located SEs.