A CONTROLLABLE GENE-EXPRESSION SYSTEM FOR THE PATHOGENIC FUNGUS CANDIDA-GLABRATA

A system for controlling gene expression was established in the pathog enic fungus Candida glabrata to elucidate the physiological functions of genes. To control the expression of the gene of interest, the C. gl abrata cells were first transformed with the plasmid carrying the tetr acycline repressor-transactivator fusion tetR::GAL4, then with the DNA fragment containing the controllable cassette, the tetracycline opera tor chimeric promoter (tetO::ScHOP1). The peptide elongation factor 3 (CgTEF3) and DNA topoisomerase II (CgTOP2) genes from C. glabrata were cloned and their expression assessed using this system. When the prom oter of CgTEF3 or CgTOP2 was replaced with tetO::ScHOP1, doxycycline a lmost completely repressed the expression of both mRNAs, and impaired growth. Repression of the TOP2 or TEF3 gene by doxycycline also hamper ed the survival of C. glabrata cells in mice; in mouse kidneys the num ber of C. glabrata cells, in which the TOP2 or TEF3 promoter was repla ced with the tetO::ScHOP1 controllable cassette, did not increase when the mice were given doxycycline. Thus, it appears that the gene repre ssion mediated by doxycycline occurred not only in culture media but a lso in animals; therefore, this system can be used to elucidate the fu nction of the gene in fungal infections and pathogenesis.