Expansion of a pivotal functional genomics resource identifies novel regulators of temperature-dependent fitness in the fungal pathogen Candida albicans

Functional genomic screens in Candida albicans are fundamental to uncover novel gene functions and the molecular mechanisms of virulence. In this study, expansion of the Gene Replacement and Conditional Expression (GRACE) library, covering now ~73% of the C. albicans genome, revealed genes relevant for fitness at cold or heat stress. From an initial screen at 30°C and 39°C, strains with moderate to severe growth defects were further assayed for growth under six different temperatures. Assessment of growth scores revealed enrichment of genes associated with RNA metabolism and regulatory-related processes as important during cold stress, whereas genes involved in cell cycle, filamentation and stress response were essential during elevated temperature. Here, we describe the function of cold-essential gene C1_11680C (or YSF3), which is involved in splicing and branchpoint recognition. RNA-Seq was conducted for functional analysis of C1_11680C, by investigating transcriptomic changes upon deletion at host (37°C) and cold stress (22°C) temperatures. This revealed major global splicing defects and confirmed its identity as the homolog of S. cerevisieae Ysf3 (Yeast splicing factor 3). We also found particular importance of this gene for splicing of introns with non-consensus branchpoints. In summary, this study describes the temperature-dependance of previously uncharazterized genes through a significant expansion of a pivotal genomics resource. We compare mRNA expression and splicing changes in different growth conditions (cold stress at 22°C, host temperature at 37° C) of Candida albicans wild-type and the GRACE strain tetO-YSF3/Δ upon induction (with Doxycycline) resulting in YSF3 depletion, and without.