In budding yeast Saccharomyces cerevisiae, Ifh1 (interacts with fork head 1) is a transcriptional co-activator that is essential for growth and is recruited to ribosomal protein gene (RPG) promoters through interaction with the forkhead-associated (FHA) domain of fork head-like 1 (Fhl1). However, the lethality of ifh1Δ cells is rescued by deletion of FHL1 or its FHA domain, suggesting that the essential function of Ifh1 is suppressing FHA domain-mediated growth inhibition. In our recent study, we found that deletion of previously uncharacterized C-terminal regions, as well as the FHA domain, of Fhl1 rescues the lethality of ifh1Δ cells. We report here that chromatin immunoprecipitation analysis of Fhl1 deletion mutants showed that deletion of the 754-844 aa region significantly decreased binding to category 1 RPG promoters, which are enriched for high mobility group 1 (Hmo1) binding, whereas deletion of the 572-662 aa region markedly reduced binding to all RPG promoters examined. These results suggest that reduced Fhl1 binding to RPG promoters enables growth of the ifh1Δ cells. Because Fhl1 binding is regulated by Hmo1 and/or FK506-sensitive proline rotamase 1 (Fpr1), we examined the effect of HMO1 deletion and found that the hmo1Δifh1Δ double mutant became viable. Yeast two-hybrid analysis revealed that the 754-844 aa region of Fhl1 interacts with Hmo1, particularly its box B domain, suggesting that this interaction contributes to the recruitment of Fhl1 to category 1 RPG promoters. Furthermore, we observed a weak interaction between Fpr1 and the 540-662 aa region of Fhl1. Genetic analysis showed that fhl1Δfpr1Δ double mutants exhibited much more severe growth defects than either single mutant, indicating that both proteins cooperatively play important roles in yeast growth, although the underlying mechanism remains unclear. Collectively, these results suggest that Hmo1 and Fpr1 regulate Fhl1 function, which exerts both negative and positive effects on yeast growth.
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