To identify physical interactors of Cas5, we leveraged the results from our immunoprecipitation coupled to mass spectrometry analysis (Supplementary Data 7). The only interaction that passed a stringent statistical analysis using Significance Analysis of Interactome (SAINT)44 with a probability threshold of 0.9 was Swi4 (Supplementary Data 7), which interacts with Swi6 to form the SBF complex in S. cerevisiae 45. To validate the interaction between Cas5 and the SBF complex, we individually tandem affinity purification (TAP) tagged Swi4 or Swi6 at the C terminus in a strain harboring Cas5-HA to enable co-immunoprecipitation analysis. The Swi4-TAP and Swi6-TAP proteins were functional and sufficient to mediate wild-type tolerance to caspofungin (Supplementary Fig. 5b). Immunoprecipitation of Cas5 with anti-HA agarose co-purified both Swi4 and Swi6 (Fig. 8a and Supplementary Fig. 6), verifying these proteins physically interact. To determine whether the SBF complex has a role in cell wall stress responses in C. albicans, we monitored the expression of ECM331 and PGA13 in mutants lacking Swi4, Swi6, or Cas5. In all three mutants, upregulation of these cell wall genes in response to caspofungin was blocked (Fig. 8b). However, strains lacking Swi4 or Swi6 displayed intermediate caspofungin susceptibility phenotypes relative to a wild-type strain and a CAS5 homozygous deletion mutant (Fig. 8c). Altogether, our results suggest that Cas5 regulates transcriptional responses to cell wall stress in part through the SBF complex and in part through an independent mechanism.Fig. 8Cas5 regulates caspofungin tolerance in part through the interaction with components of the SBF complex, Swi4 and Swi6. a Swi4 and Swi6 co-purify with Cas5. C terminally HA-tagged Cas5 was immunoprecipitated with α-HA beads. Swi4 and Swi6 co-purification was monitored by western blot and detected with an α-TAP antibody. Cas5 pull down was confirmed by detection with an α-HA antibody. Input samples confirm the expression of tagged proteins. Full blots are shown in Supplementary Fig. 6. b Cas5, Swi4, and Swi6 are required for the upregulation of caspofungin-responsive genes. The transcript levels of ECM331 and PGA13 was monitored by qRT-PCR and normalized to GPD1. Plotted are the fold-changes in transcript levels in the presence of caspofungin relative to untreated. Error bars represent standard deviation (s.d.) from the mean of triplicate samples. The fold change in gene expression for each mutant strain was compared to the wildtype using one-way ANOVA in GraphPad Prism (****P < 0.0001). c Swi4 and Swi6 regulate cell wall stress response. Caspofungin susceptibility assays were conducted in YPD medium. Growth was measured by absorbance at 600 nm after 48 h at 30 °C. Optical densities were averaged for duplicate measurements. Data was quantitatively displayed with color using Treeview (see color bar in Fig. 2)