Rapid movement of newly synthesized chicken apolipoprotein AI to trans-Golgi network and its secretion in Madin-Darby canine kidney cells.
MDCK cells were utilized to study the biosynthesis and secretion of chicken apolipoprotein AI (apoAI). A full-length apoAI cDNA in an eukaryotic expression vector was transfected into a MDCK-TGG cell line, expressing a trans-Golgi network (TGN) marker (TGG), and stable clones expressing apoAI were selected. Pulse-chase and cell fractionation studies showed that, compared to gp 80 (an endogenous secretory protein), apoAI was rapidly transported from RER to Golgi complex within 5 min and released from the Golgi complex to the cell exterior by 30 min. Immunofluorescence and three-dimensional laser scanning confocal microscopy revealed that at steady state apoAI was predominantly localized in the TGN. Transferrin uptake experiments showed that apoAI, localized in the TGN, was derived primarily from biosynthetic and not from endocytic routes. ApoAI was secreted in a nonpolarized manner. We suggest that apoAI stays transiently in the TGN prior to its secretion and that the major events of apoAI biosynthesis in vivo and in MDCK cells are conserved. Possible mechanisms of rapid ER to Golgi transport and TGN localization of apoAI are discussed.
Banerjee
D
D
The Lindsley F. Kimball Research Institute of The New York Blood Center, New York 10021, USA.
Rodriguez
M
M
Rajasekaran
A K
AK
D000818:Animals; D016632:Apolipoprotein A-I / Q000096:biosynthesis* / Q000378:metabolism*; D001692:Biological Transport; D020126:Brefeldin A; D002460:Cell Line; D016764:Cell Polarity; D002645:Chickens; D003517:Cyclopentanes / Q000494:pharmacology; D004285:Dogs; D018870:Endoplasmic Reticulum, Rough / Q000378:metabolism; D004847:Epithelial Cells; D006056:Golgi Apparatus / Q000378:metabolism*; D006596:Hexosaminidases; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology
D016632:Apolipoprotein A-I; D003517:Cyclopentanes; D011500:Protein Synthesis Inhibitors; D020126:Brefeldin A; D006596:Hexosaminidases