Selective modulation of P-glycoprotein's ATPase and anion efflux regulation activities with PKC alpha and PKC epsilon in Sf9 cells.
The modulation of P-glycoprotein's (Pgp) ATPase activity and its ability to regulate swelling-activated 125I efflux, by PKC alpha and PKC epsilon, was examined in insect cells. Recombinant baculovirus was used to express human Pgp in Sf9 cells and Pgp was also coexpressed with either PKC alpha or PKC epsilon. ATPase assays showed the enzyme activity of Pgp to be elevated during co-expression with the Ca2+ dependent isoform PKC alpha, but not with the Ca2+ independent variant PKC epsilon. Furthermore, neither isoform, when co-expressed with Pgp, altered the swelling-activated efflux of 125I from Sf9 cells. However, in cells co-expressing Pgp/PKC (alpha or epsilon), pre-treatment with the phorbol ester TPA significantly reduced the swelling-activated 125I efflux with both PKC isoforms. Our results suggest that phosphorylation with the Ca2+ independent variant PKC epsilon does not regulate the ATPase activity of Pgp and that stimulation of PKC with TPA alters the swelling-activated efflux of anions from insect cells expressing Pgp.
Idriss
H
H
Nuffield Dept. of Clinical Biochemistry, Oxford University, Headington, UK. hi@st-and.ac.uk
Urquidi
V
V
Basavappa
S
S
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism*; D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000818:Animals; D000838:Anions / Q000378:metabolism; D016367:Baculoviridae / Q000235:genetics; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D004586:Electrophoresis; D006801:Humans; D007120:Immunochemistry; D007313:Insecta / Q000378:metabolism; D007457:Iodine Radioisotopes; D007527:Isoenzymes / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D051571:Protein Kinase C-alpha
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000838:Anions; D007457:Iodine Radioisotopes; D007527:Isoenzymes; C115197:calcium-independent protein kinase C; C496628:PRKCA protein, human; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D000251:Adenosine Triphosphatases