Evolutionary rewiring of the ROX genes controlling hypoxic response in yeasts

Unusually among fungi, Saccharomyces cerevisiae is able to grow in environments containing almost no oxygen. A major feature of its response to hypoxia is a transition in expression from aerobic to hypoxic genes, which often code for duplicated isoforms of the same protein. In aerobic conditions, expression of the hypoxic gene set is repressed by the HMG domain protein Rox1. Here, we examined the evolution of ROX1 and related genes in the subphylum Saccharomycotina and find that a substantial reorganization of hypoxic gene regulation occurred during yeast evolution. S. cerevisiae lost ROX2, an ancient paralog of ROX1, which is almost universally present in other yeast species. ROX2 is orthologous to Candida albicans RFG1, a regulator of filamentous growth. Many yeasts, such as Candida glabrata, lack ROX1 and contain only ROX2. Others such as Naumovozyma castellii retain both genes. Although the ancestral function of ROX2 is uncertain, we find that it is not a regulator of hypoxic genes except in C. glabrata where it has taken over this function from the absent ROX1. We also find that N. castellii has a greatly attenuated transcriptional response to hypoxia as compared to other species, but that the ergosterol pathway which is normally induced by hypoxia can be induced by cobalt chloride stress in N. castellii. Overall design: N. castellii mRNA profiles of three biological replicates of wild type (WT) and rox2-/-, and two biological replicates of rox1-/-, rox1/rox2-/-, WT + CoCl2, rox1-/- + CoCl2, rox2-/- + CoCl2 and rox1/rox2-/- + CoCl2 were generated by deep sequencing, using Illumina GAII X. N. castellii mRNA profiles of WT in normoxic and hypoxic conditions with three biological replicates each were generated by BGI using Illumina HiSeq 2000.