Inhibition of N-linked glycosylation of P-glycoprotein by tunicamycin results in a reduced multidrug resistance phenotype.
Characterisation of altered glycosylation of P-glycoprotein (P-gp) found associated with the absence of a multidrug resistance (MDR) phenotype in cell lines prompted an investigation to assess the role of post-translational processing in establishing P-gp efflux pump functionally. The clone A cell line used in this study displays a strong MDR phenotype mediated by high constitutive levels of expression of P-gp. Incubation of clone A cells with tunicamycin for different periods resulted in a time-dependent increase in daunorubicin accumulation, reflecting a reduction in P-gp function. Parallel experiments conducted with verapamil resulted in no loss of P-gp functionality in clone A cells. Reduction in surface-associated P-gp following exposure to tunicamycin was established by FACS analysis, Western blot analysis and immunoprecipitation of surface-iodinated P-gp. In addition, immunoprecipitation of P-gp from 32P-orthophosphate-labelled cells demonstrated reduced phosphorylation of P-gp associated with tunicamycin exposure. From these studies we conclude that glycosylation of P-gp is required to establish the cellular MDR phenotype.
Kramer
R
R
American Cyanamid Company, Medical Research Division, Pearl River, NY 10965, USA.
Weber
T K
TK
Arceci
R
R
Ramchurren
N
N
Kastrinakis
W V
WV
Steele
G
G
Summerhayes
I C
IC
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D015153:Blotting, Western; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D015179:Colorectal Neoplasms; D018432:Drug Resistance, Multiple; D066246:ErbB Receptors / Q000302:isolation & purification / Q000378:metabolism; D005434:Flow Cytometry; D006031:Glycosylation / Q000187:drug effects; D006801:Humans; D010641:Phenotype; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational / Q000187:drug effects*; D014407:Tumor Cells, Cultured; D014415:Tunicamycin / Q000494:pharmacology*
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D014415:Tunicamycin; D066246:ErbB Receptors