Characterization of the rebinding of 125I-epidermal growth factor released from BALB/c-3T3 cells following accumulation in the presence of chloroquine.
Lysosomotropic amines, such as chloroquine and methylamine, increase the intracellular accumulation of 125I-EGF by inhibiting lysosomal degradation. It has been shown previously that BALB/c-3T3 cells, prelabeled at 4 degrees C with 125I-EGF for 3 h and subsequently chased at 37 degrees C in the presence of chloroquine, internalized the surface bound 125I-EGF which was subsequently released into the extracellular medium in a high molecular weight form which co-migrated with native 125I-EGF. The secreted 125I-EGF rebound to the cells from which it was released more efficiently than does peptide in the extracellular media. We now show that when the BALB/c-3T3 cells were prelabeled at 37 degrees C for 2 h in the presence of chloroquine, the internalized 125I-EGF released into the medium was in a high molecular weight form which co-migrated with native 125I-EGF and did not rebind anymore efficiently than did peptide in the extracellular media. This lack of rebinding was not due to an alteration in the 125I-EGF molecule since it was still capable of rebinding to naive A431 cells, nor was it due to the exhaustion of EGF receptors on the BALB/c-3T3 cells. The inhibition of rebinding was observed only when the cells were treated with EGF in the presence of chloroquine, and was not due to a general down-regulation of membrane receptors. The differences between the rebinding of 125I-EGF at 4 degrees C and 37 degrees C suggest that EGF may be processed via different pathways in the cell.
Cooper
J L
JL
University of California, Los Alamos National Laboratory, New Mexico 87545.
Wakshull
E
E
Wharton
W
W
D000818:Animals; D002460:Cell Line; D002738:Chloroquine / Q000494:pharmacology*; D003470:Culture Media; D004815:Epidermal Growth Factor / Q000378:metabolism*; D066246:ErbB Receptors / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008970:Molecular Weight; D013696:Temperature; D013997:Time Factors
D003470:Culture Media; D004815:Epidermal Growth Factor; D002738:Chloroquine; D066246:ErbB Receptors