Golgi retardation in Madin-Darby canine kidney and Chinese hamster ovary cells of a transmembrane chimera of two surface proteins.
Dipeptidyl peptidase IV (DDD) is a type II plasma membrane protein. Replacement of its transmembrane domain with that of another surface protein, aminopeptidase N, resulted in accumulation in the Golgi apparatus of Madin-Darby canine kidney cells and a delayed Golgi to surface transport in Chinese hamster ovary (CHO) cells. The compartment of retardation was identified as post medial-Golgi, most likely to be the trans-Golgi/trans Golgi network (TGN). Compared to native DDD, the rate of endoplasmic reticulum to Golgi transport for the chimera was largely unchanged in both cell types. On the other hand, Golgi to surface transport was delayed by more than 2 h in CHO cells and essentially undetectable up to 22 h of chase in Madin-Darby canine kidney cells. The decrease in the rate of Golgi to surface transport in CHO cells resulted in a significant accumulation of the fusion protein in the trans-Golgi/TGN. This phenomena is very unlikely to be due to any drastic conformational changes, as neither the enzyme activity nor the dimerization of the constructed molecule was affected. The findings of this study indicate that the transmembrane domain, in the context of its flanking sequences, is important for efficient Golgi to cell surface transport.
Low
S H
SH
Membrane Biology Laboratory, National University of Singapore.
Tang
B L
BL
Wong
S H
SH
Hong
W
W
D000595:Amino Acid Sequence; D000626:Aminopeptidases / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D020126:Brefeldin A; D018826:CD13 Antigens; D016466:CHO Cells; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology*; D002846:Chromatography, Affinity; D006224:Cricetinae; D003517:Cyclopentanes / Q000494:pharmacology; D017931:DNA Primers; D018819:Dipeptidyl Peptidase 4; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D004285:Dogs; D004591:Electrophoresis, Polyacrylamide Gel; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D006056:Golgi Apparatus / Q000378:metabolism*; D007668:Kidney; D007700:Kinetics; D037102:Lectins; D008565:Membrane Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D013347:Subcellular Fractions / Q000187:drug effects / Q000378:metabolism
D003517:Cyclopentanes; D017931:DNA Primers; D037102:Lectins; D008565:Membrane Proteins; D011500:Protein Synthesis Inhibitors; D011993:Recombinant Fusion Proteins; D020126:Brefeldin A; D000626:Aminopeptidases; D018826:CD13 Antigens; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases; D018819:Dipeptidyl Peptidase 4