Multidrug resistance after retroviral transfer of the human MDR1 gene correlates with P-glycoprotein density in the plasma membrane and is not affected by cytotoxic selection.
Multidrug resistance (MDR) in mammalian cells is associated with the expression of the MDR1 gene encoding P-glycoprotein (P-gp), an and active efflux pump for various lipophilic compounds. MDR transfectants can be isolated after MDR1 gene transfer and selection with cytotoxic drugs; low levels of drug resistance have also been observed in unselected NIH 3T3 mouse cells after retrovirus-mediated transfer of mouse mdr1 cDNA. MDR cell lines possess multiple phenotypic changes, suggesting that P-gp function could be complemented by some additional mechanisms associated with cytotoxic selection. To determine whether cytotoxic selection contributes to the MDR phenotype of MDR1-expressing cells, NIH 3T3 cells infected with a recombinant retrovirus carrying the human MDR1 gene were selected by two different procedures: (i) noncytotoxic selection for increased P-gp expression on the cell surface by multiple rounds of immunofluorescence labeling and flow sorting or (ii) one or more steps of selection with a cytotoxic drug. The levels of MDR in both types of infectants showed an excellent correlation with the P-gp density in the plasma membrane, expressed as immunoreactivity with a P-gp-specific antibody normalized by reactivity with an antibody against an unrelated antigen. Cytotoxic selection conferred no additional increase in resistance relative to P-gp density. These results indicate that P-gp density in the plasma membrane may be sufficient to determine the level of MDR.
Department of Genetics, University of Illinois, Chicago 60612.
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K
Choi
T O
TO
Frommel
R K
RK
Stern
C F
CF
Perez
M
M
Kriegler
T
T
Tsuruo
I B
IB
Roninson
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000818:Animals; D015139:Blotting, Southern; D002460:Cell Line; D002462:Cell Membrane / Q000502:physiology; D002470:Cell Survival / Q000187:drug effects; D004351:Drug Resistance / Q000235:genetics*; D005455:Fluorescent Antibody Technique; D006801:Humans; D008562:Membrane Glycoproteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D012190:Retroviridae / Q000235:genetics; D014162:Transfection; D014747:Vinblastine / Q000494:pharmacology*
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; D014747:Vinblastine