The Candida albicans reference strain SC5314 contains a rare, dominant allele of the transcription factor Rob1 that modulates biofilm formation and oral commensalism

Candida albicans is a diploid human fungal pathogen that displays significant genomic and phenotypic heterogeneity over a range of virulence traits and in the context of a variety of environmental niches. Here, we show that the effects of Rob1 on biofilm and filamentation virulence traits is dependent on both the specific environmental condition and the clinical strain of C. albicans. The C. albicans reference strain SC5314 is a ROB1 heterozygote with two alleles that differ by a single nucleotide polymorphism at position 946 resulting in a serine or proline containing isoform. An analysis of 224 sequenced C. albicans genomes indicates that SC5314 is the only ROB1 heterozygote documented to date and that the dominant allele contains a proline at position 946. Remarkably, the ROB1 alleles are functionally distinct and the rare ROB1946S allele supports increased filamentation in vitro and increased biofilm formation in vitro and in vivo, suggesting it is a phenotypic gain-of-function allele. SC5314 is amongst the most highly filamentous and invasive strains characterized to date. Introduction of the ROB1946S allele into a poorly filamenting clinical isolate increases filamentation and conversion of an SC5314 laboratory strain to a ROB1946S homozygote increases in vitro filamentation and biofilm formation. In a mouse model of oropharyngeal infection, the predominant ROB1946P allele establishes a commensal state while the ROB1946S phenocopies the parent strain and invades into the mucosae. These observations provide an explanation for the distinct phenotypes of SC5314 and highlight the role of heterozygosity as a driver of C. albicans phenotypic heterogeneity. Overall design: An RNAseq approach was used to investigate the role of Rob1 in the regulation of gene expression during in vitro hyphal induction conditions. C. albicans strain SN250 (WT) or the same strain with both alleles of Rob1 deleted (ROB1NULL) were grown for 4 hours under three different conditions. Growth at 37C in RPMI media or SPIDER media were used to induce hyphal growth. The third control condition was growth at 30C in YPD media. Each condition was performed in duplicate (A and B). Differential expresssion analysis was performed on the RNAseq gene count data to identify Rob1-dependent gene expression in each of the growth conditions.