Candida glabrata ATP-binding cassette transporters Cdr1p and Pdh1p expressed in a Saccharomyces cerevisiae strain deficient in membrane transporters show phosphorylation-dependent pumping properties.
The expression and drug efflux activity of the ATP binding cassette transporters Cdr1p and Pdh1p are thought to have contributed to the recent increase in the number of fungal infections caused by Candida glabrata. The function of these transporters and their pumping characteristics, however, remain ill defined. We have evaluated the function of Cdr1p and Pdh1p through their heterologous hyperexpression in a Saccharomyces cerevisiae strain deleted in seven major drug efflux transporters to minimize the background drug efflux activity. Although both Cdr1p- and Pdh1p-expressing strains CDR1-AD and PDH1-AD acquired multiple resistances to structurally unrelated compounds, CDR1-AD showed, in most cases, higher levels of resistance than PDH1-AD. CDR1-AD also showed greater rhodamine 6G efflux and resistance to pump inhibitors, although plasma membrane fractions had comparable NTPase activities. These results indicate that Cdr1p makes a larger contribution than Phd1p to the reduced susceptibility of C. glabrata to xenobiotics. Both pump proteins were phosphorylated in a glucose-dependent manner. Whereas the phosphorylation of Cdr1p affected its NTPase activity, the protein kinase A-mediated phosphorylation of Pdh1p, which was necessary for drug efflux, did not. This suggests that phosphorylation of Pdh1p may be required for efficient coupling of NTPase activity with drug efflux.
Wada
Shun-Ichi
S
Department of Bioactive Molecules, National Institute of Infectious Diseases, 1-23-1 Toyama, Shinjuku-ku, Tokyo 162-8640, Japan.
Niimi
Masakazu
M
Niimi
Kyoko
K
Holmes
Ann R
AR
Monk
Brian C
BC
Cannon
Richard D
RD
Uehara
Yoshimasa
Y
D018528:ATP-Binding Cassette Transporters / Q000037:antagonists & inhibitors; D000595:Amino Acid Sequence; D000935:Antifungal Agents / Q000494:pharmacology; D001483:Base Sequence; D001692:Biological Transport; D015152:Blotting, Northern; D041221:Candida glabrata / Q000378:metabolism*; D017931:DNA Primers; D015725:Fluconazole / Q000494:pharmacology; D005656:Fungal Proteins; D008565:Membrane Proteins / Q000235:genetics*; D026901:Membrane Transport Proteins / Q000235:genetics*; D008826:Microbial Sensitivity Tests; D010766:Phosphorylation; D012333:RNA, Messenger / Q000235:genetics; D012235:Rhodamines / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins
D018528:ATP-Binding Cassette Transporters; D000935:Antifungal Agents; C094693:CDR1 protein, Candida albicans; D017931:DNA Primers; D005656:Fungal Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D012333:RNA, Messenger; D012235:Rhodamines; D029701:Saccharomyces cerevisiae Proteins; C026188:rhodamine 6G; D015725:Fluconazole