Effect of CCH1 and MID1 in calcium influx under alkaline pH and its regulation by Crz1p transcription factor in Candida albicans

Abstract:

In Candida albicans, adaptation to environmental pH is relevant to its pathogenicity. Calcium signaling pathway involves in many stress responses and often accompany with Ca2+ fluctuation. We constructed CCH1 and MID1 mutant strains and studied their effect on calcium influx and further investigated the regulation by Crz1p transcription factor. We used PCR-directed gene disruption to construct cch1Δ/Δ and mid1Δ/Δ null mutant. By using a flow cytometry-based method we monitored the free cytosolic Ca2+ levels under alkaline stress. Moreover, we constructed pPHO89-LacZ plasmids and by β-Galactosidase assays, we analyzed the changes of LacZ activities after gene disruption. The results showed that alkaline stress induced calcium burst reduced obviously in cch1Δ/Δ and mid1Δ/Δ mutant strains, also for LacZ activities, and fully abolished in crz1Δ/Δ mutant strain. Finally, by realtime PCR, we confirmed the regulation role of Crz1p in CCH1 and MID1 genes but in a calcineurin independent way. Studies on the effect of calcium pathway on response to alkaline stress will provide an important theoretical basis for Candida albicans infection-oriented treatment and new drug targets.

Key words:Candida albicans, alkaline pH, CCH1, MID, calcium influx, Crz1p