Study On Redox Function Of Novel Gene Ipf19998 In Candida Albicans

More recently, it has been widely accepted that Candida albicans can exploit several cellular responses to facilitate tolerance of antifungal agents and can further acquire resistance by multiple mechanisms. Now the sequencing results of the Candida albicans genome has presented and could be gained in public database. This study finds that a novel gene IPF19998 in Candida albicans shows highly homology with gene AIF1 in Saccharomyces cerevisiae after blastn and blastp. It has proved that in Saccharomyces cerevisiae Aif1p located in mitochondria and translocates to the nucleus of yeast cells in response to apoptotic stimuli like oxygen stress age-induced apoptosis[1]. So we presumed that the orthologue of IPF19998 in Candida albicans showed the same localization and exhibits similar death executing pathways as yeast AIF1. It may play an important role for explaining the mechanism of apoptosis in Candida albicans and apply for a new way for anti-fungal drugs researching.We constructed the IPF19998 gene knock-out strain with Ura-Blaster strategies, and verificated by PCR and southern blotting and also the ectopic overexpression of IPF19998 gene in the ipf19998 mutants, verificated by real-time PCR. Then, growth curve and microdilution test were investigated in determination of susceptibility of C. albicans treated by antifungals and H2O2. Cultivate the strains following the formula of the Candida albicans biofilms for 24 hours. The endogenous ROS production for both biofilm and wild type were tested by fluorochrome. Moreover, any point mutations in other known oxidative stress-related genes and biofilm forming-related genes are further examined in IPF19998 null mutant. Conclusions: disrupted the two alleles of IPF19998 sequentially in CAI4 strain using the URA-blaster method and 5-FOA selection, constructed the overexpression-gene strain successfully. We demonstrated that after disruption of the IPF19998 gene, it had no differences in speed of strain growth and the sensitivity to azole-drugs or H2O2 comparing with wild type strain. But, the overexpressed strain showed resistance to azole-drugs. The mutant showed weak ability to form the biofilm and higher endogenous ROS production in biofilm while the overexpression strain could form a strong one. It was verificated also in realtime PCR. The knock-out strain showed different expression of the known oxidative stress-related genes and biofilm forming-related genes. The results indicats that product encoded by IPF19998 gene may be involved in the anti-oxidative capacities in biofilm-forming but not apoptosis we expected before. On the whole, a great of experiments are still needed to illustrate the definite role played by IPF19998 in the pathway.