Investigation on MFS family in C. parapsilosis with CRISPR

C. parapsilosis causes candidiasis especially among newborn babies. The function of specific transporters is considered to be one key feature underlying drug resistance in Candida species. Drug transporters fall into two main classes – ATP-binding cassette (ABC) transporters, and the major facilitator superfamily (MFS). Particularly, Some members of members of the drug/H (+) antiporter family (DHA1) of the MFS superfamily function as multidrug transporters. We find that the DHA1 family in Candida species can be divided into several clades. These include MDR1/FLR1, associated with multidrug resistance in C. albicans (1 member in C. parapsilosis); TPO4, associated with polyamine transport (1 member in C. parapsilosis); NAG3/4, associated with transport of N-acetyl glucosamine (2 members in C. parapsilosis); TPO2/3, associated with polyamine transport (1 member in C. parapsilosis); YHR048w, with no known function (no members in C. parapsilosis); and TPO1/FLU1, possibly associated with fluconazole resistance (8 members in C. parapsilosis). We propose to use CRISPR-based gene editing to explore the function of all 13 members of the DHA1 family in C. parapsilosis. To date we have individually edited 10 members of the family by introducing stop codons near the start site of translation (ATG). We are currently editing the remaining 3 genes, and we are attempting to combine at least two edited genes in the same background. We will then test the phenotype of each edited strain in the presence of various drugs.