Intracellular trafficking of recycling apolipoprotein E in Chinese hamster ovary cells.
We have investigated apolipoprotein E (apoE) recycling in Chinese hamster ovary (CHO) cells, a peripheral cell that does not produce lipoproteins or express apoE. Using a pulse-chase protocol in which cells were pulsed with 125I-apoE-VLDL and chased for different periods, approximately 30% of the apoE internalized during the pulse was resecreted within a 4 h chase in a relatively lipid-free state. The addition of lysosomotropic agents or brefeldin A had no effect on apoE recycling. Unlike previous results with hepatocytes and macrophages, neither apoA-I nor upregulation of ABCA1 stimulated apoE recycling. However, cyclodextrin, which extracts cholesterol from plasma membrane lipid rafts, increased recycling. Confocal studies revealed that apoE, internalized during a 1 h pulse, colocalizes with early endosomal antigen-1, Rab5, Rab11a, and lysobisphosphatidic acid but not with lysosomal-associated membrane protein-1. Colocalization of apoE and Rab11a persisted even after cells had been chased for 1 h, suggesting a pool of apoE within the endosomal recycling compartment (ERC). Our data suggest that apoE recycling in CHO cells is linked to cellular cholesterol removal via the ERC and phospholipid-containing acceptors in a pathway alternative to the ABCA1-apoA-I axis.
Department of Pathology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA.
Nicole A
NA
Braun
Peter J
PJ
Mohler
Karl H
KH
Weisgraber
Alyssa H
AH
Hasty
MacRae F
MF
Linton
Patricia G
PG
Yancey
Yan Ru
YR
Su
Sergio
S
Fazio
Larry L
LL
Swift
D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters / Q000378:metabolism / Q000502:physiology; D000818:Animals; D016632:Apolipoprotein A-I / Q000378:metabolism / Q000502:physiology; D001057:Apolipoproteins E / Q000378:metabolism*; D020126:Brefeldin A / Q000494:pharmacology; D016466:CHO Cells; D002738:Chloroquine / Q000494:pharmacology; D002784:Cholesterol / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D003505:Cyclodextrins / Q000494:pharmacology; D005260:Female; D005455:Fluorescent Antibody Technique; D006801:Humans; D008079:Lipoproteins, VLDL / Q000378:metabolism*; D018613:Microscopy, Confocal; D008985:Monensin / Q000494:pharmacology; D021381:Protein Transport / Q000187:drug effects
C577836:ABCA1 protein, human; D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters; D016632:Apolipoprotein A-I; D001057:Apolipoproteins E; D003505:Cyclodextrins; D008079:Lipoproteins, VLDL; D020126:Brefeldin A; D002738:Chloroquine; D008985:Monensin; D002784:Cholesterol