Wortmannin alters the transferrin receptor endocytic pathway in vivo and in vitro.
Treatment with the phosphatidylinositol 3-kinase inhibitor wortmannin promotes approximately 30% decrease in the steady-state number of cell-surface transferrin receptors. This effect is rapid and dose dependent, with maximal down-regulation elicited with 30 min of treatment and with an IC50 approximately 25 nM wortmannin. Wortmannin-treated cells display an increased endocytic rate constant for transferrin internalization and decreased exocytic rate constants for transferrin recycling. In addition to these effects in vivo, wortmannin is a potent inhibitor (IC50 approximately 15 nM) of a cell-free assay that detects the delivery of endocytosed probes into a common compartment. Inhibition of the in vitro assay involves the inactivation of a membrane-associated factor that can be recruited onto the surface of vesicles from the cytosol. Its effects on the cell-free assay suggest that wortmannin inhibits receptor sorting and/or vesicle budding required for delivery of endocytosed material to "mixing" endosomes. This idea is consistent with morphological changes induced by wortmannin, which include the formation of enlarged transferrin-containing structures and the disruption of the perinuclear endosomal compartment. However, the differential effects of wortmannin, specifically increased transferrin receptor internalization and inhibition of receptor recycling, implicate a role for phosphatidylinositol 3-kinase activity in multiple sorting events in the transferrin receptor's membrane traffic pathway.
Spiro
D J
DJ
Program in Biological and Biomedical Sciences, Harvard Medical School, Boston, Massachusetts 02115, USA.
Boll
W
W
Kirchhausen
T
T
Wessling-Resnick
M
M
D000730:Androstadienes / Q000494:pharmacology*; D015536:Down-Regulation; D004705:Endocytosis; D011992:Endosomes; D006367:HeLa Cells; D006801:Humans; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors*; D011990:Receptors, Transferrin / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D000077191:Wortmannin
D000730:Androstadienes; D011990:Receptors, Transferrin; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D000077191:Wortmannin