Fig 6. Mfg1 and Flo8 bind to the promoter of TEC1 to regulate its expression, and overexpression of FLO8 drives TEC1 expression, enabling filamentation in the absence of Mfg1.

A) Overexpression of TEC1 results in filamentous growth in the absence of MFG1 or FLO8. Cells were grown in YPD at 30°C or in YPD with 10% serum at 37°C for 5 hours. Scale bar is 20 μm. B) Binding of Flo8-TAP and Mfg1-TAP to the promoter of TEC1 was assessed using ChIP-qPCR. Shown is the fold-enrichment over the untagged parental strain, which is set at 1. Asterisks indicate P < 0.01 (**) or P < 0.05 (*), relative to the wild-type parent in the respective condition (two-tailed unpaired t-test). Error bars represent the standard deviation of technical triplicates. Assays were performed in biological duplicate. C) TEC1 expression is reduced in the absence of Flo8 or Mfg1. Cells were grown in YPD at 30°C for 3.5 hours, pelleted and washed, and then transferred to YPD at 30°C or to YPD at 37°C with 10% serum for 1 hour. Transcript levels were monitored using qRT-PCR and normalized to PMA1 and RIP1. Error bars represent standard error of technical triplicates. Assays were performed in biological duplicate. Asterisks indicate P < 0.001 (***), relative to the wild-type strain in each respective condition (two-way ANOVA, Bonferroni Multiple Comparison Test). D) TEC1 expression is increased in strains overexpressing FLO8. Cells were grown in YPD at 30°C for 4.5 hours. Transcript levels were monitored using qRT-PCR and normalized to PMA1 and RIP1. Error bars represent standard error of technical triplicates. Assays were performed in biological duplicate. Asterisks indicate P < 0.0001 (***), relative to the respective parental strain, or as indicated (one-way ANOVA, Bonferroni Multiple Comparison Test).