Chemiluminescence of superoxide generated by Candida albicans: differential effects of the superoxide generator paraquat on a wild-type strain and a respiratory mutant.
We previously reported that a respiration-competent parent strain (K) of Candida albicans was more susceptible to the intracellular superoxide radical (O2-) generator paraquat (PQ) than was a respiration-deficient mutant (KRD-19), although both showed a similar sensitivity to extracellularly generated O2-. To clarify the cause of the differential PQ lethality, we developed a chemiluminescence method for measuring O2- generated by C. albicans cells by using the probe methyl-Cypridina-luciferin analogue (MCLA), and examined the effects of PQ on O2- generation in both parent and mutant strains. Endogenous O2- generation without stimulation by PQ was unexpectedly low in both strains. PQ-induced O2- generation in the parent strain was maximal in logarithmic phase cells and lowered in stationary phase cells. In contrast, O2- generation in the mutant remained low throughout the growth phase, even when stimulated by PQ. The extent of PQ-induced O2- generation in the parent strain depended on the carbon source added to the assay mixture; in decreasing order, glucose, glycerol, no carbon source. The inhibitor of the cytochrome respiratory chain, antimycin A, suppressed almost completely the PQ-induced O2- generation in the parent strain. It has been established that PQ is converted to its radical form (PQ+) by receiving a single electron in cells. PQ+ then reduces molecular oxygen to O2- by redox cycling. Thus, the high tolerance to PQ of the respiration-deficient mutant can be explained by minimal PQ+/O2- production due to the limited supply of electrons from the impaired respiratory system.
Aoki
S
S
Advanced Research Center, Nippon Dental University, Niigata, Japan. aokis@ngt.ndu.ac.jp
Ito-Kuwa
S
S
Nakamura
K
K
Nakamura
Y
Y
Vidotto
V
V
Takeo
K
K
D002176:Candida albicans / Q000187:drug effects* / Q000254:growth & development / Q000378:metabolism; D002452:Cell Count; D007093:Imidazoles / Q000494:pharmacology; D008163:Luminescent Measurements; D009154:Mutation; D010101:Oxygen Consumption; D010269:Paraquat / Q000494:pharmacology*; D011719:Pyrazines / Q000494:pharmacology; D013481:Superoxides / Q000378:metabolism*
D007093:Imidazoles; D011719:Pyrazines; D013481:Superoxides; C063412:2-methyl-6-(4-methoxyphenyl)-3,7-dihydroimidazo(1,2-alpha)pyrazin-3-one; D010269:Paraquat