Clustered folate receptors deliver 5-methyltetrahydrofolate to cytoplasm of MA104 cells.
Previously, a high affinity, glycosylphosphatidylinositol-anchored receptor for folate and a caveolae internalization cycle have been found necessary for potocytosis of 5-methyltetrahydrofolate in MA104. We now show by cell fractionation that folate receptors also must be clustered in caveolae for potocytosis. An enriched fraction of caveolae from control cells retained 65-70% of the [3H]folic acid bound to cells in culture. Exposure of cells to the cholesterol-binding drug, filipin, which is known to uncluster receptors, shifted approximately 50% of the bound [3H]folic acid from the caveolae fraction to the noncaveolae membrane fraction and markedly inhibited internalization of [3H]folic acid. An mAb directed against the folate receptor also shifted approximately 50% of the caveolae-associated [3H]folic acid to noncaveolae membrane, indicating the antibody perturbs the normal receptor distribution. Concordantly, the mAb inhibited the delivery of 5-methyl[3H]tetrahydrofolate to the cytoplasm. Receptor bound 5-methyl[3H]tetrahydrofolate moved directly from caveolae to the cytoplasm and was not blocked by phenylarsine oxide, an inhibitor of receptor-mediated endocytosis. These results suggest cell fractionation can be used to study the uptake of molecules by caveolae.
Smart
E J
EJ
Department of Cell Biology and Neuroscience, University of Texas Southwestern Medical Center, Dallas 75235-9039, USA.
Mineo
C
C
Anderson
R G
RG
D000818:Animals; D001692:Biological Transport; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D003593:Cytoplasm / Q000378:metabolism; D058973:Folate Receptors, GPI-Anchored; D005492:Folic Acid / Q000378:metabolism; D000882:Haplorhini; D011940:Receptor Aggregation; D011956:Receptors, Cell Surface / Q000378:metabolism*; D013763:Tetrahydrofolates / Q000378:metabolism*
D002352:Carrier Proteins; D058973:Folate Receptors, GPI-Anchored; D011956:Receptors, Cell Surface; D013763:Tetrahydrofolates; D005492:Folic Acid; C005984:5-methyltetrahydrofolate