, 2000, 2001). The N-terminal domain of Bcy1 served to target it properly during logarithmic and stationary phase (Griffioen et al., 2000). Phosphorylation of its N-terminal domain directed Bcy1 to cytoplasm. Bcy1 modification was found to be dependent on Yak1 kinase (Griffioen et al., 2001). Zds1-mediated cytoplasmic localization
Trametinib of Bcy1 was regulated by carbon source-dependent phosphorylation of cluster II serines (Griffioen et al., 2001; Griffioen & Thevelein, 2002). Recently, we reported that Sch9 was involved in regulating phosphorylation and localization of Bcy1 (Zhang et al., 2011). But the mechanisms of Sch9 regulating Bcy1 are still unknown. The serine/threonine protein kinase, Yak1, functioned as a negative regulator of the cell cycle in S. cerevisiae, acting downstream of the cAMP-dependent protein kinase (Garrett & Broach, 1989). Yak1 is a dual specificity Transmembrane Transporters modulator protein kinase which autophosphorylates on Tyr-530 and phosphorylates exogenous substrates on
Ser/Thr residues (Kassis et al., 2000). When glucose is limited, Yak1 accumulates in the nucleus where it phosphorylates Pop2, which is required for proper cell cycle arrest. In the presence of glucose, Yak1 was phosphorylated by an as yet unknown protein kinase at its serine residue(s) and associates with Bmh1 and Bmh2, and was then exported from the nucleus to the cytoplasm (Moriya et al., 2001). ZDS1 and ZDS2 of S. cerevisiae were reported to be involved in transcriptional silencing, longevity, optimal mRNA export and mitotic exit through regulation of Cdc14 (Roy & Runge, 2000; Estruch et al., 2005; Queralt & Uhlmann, 2008). Zds1 was also reported to control sexual differentiation, cell wall integrity and cell morphology in fission yeast (Yakura et al., 2006). Recently, it was reported that that Zds1/Zds2 primarily control localization of Cdc55, a regulatory B subunit of the PP2A, which plays important roles in mitotic entry and mitotic exit (Rossio & Yoshida, 2011). Here we report that Sch9 regulates the localization of Bcy1 via Zds1 by showing that: (1) deletion of SCH9 or ZDS1 both
caused nuclear localization of Bcy1; (2) Sch9 and Zds1 interacted physically; (3) overexpression of ZDS1 led to a significant increased cytoplasmic localization of Bcy1 in sch9Δ cells, whereas PD184352 (CI-1040) overexpression of SCH9 had no visible effect on cytoplasmic localization of Bcy1 in zds1Δ cells. Additionally, our study suggests that Sch9 regulated the phosphorylation of Bcy1 via Yak1. Yeast cells were grown in YPD [1% (w/v) yeast extract, 2% (w/v) peptone and 2% (w/v) glucose] or in synthetic complete (SC) medium [0.17% (w/v) nitrogen base, with adenine, uracil, histidine, leucine, tryptophan and amino acids as appropriate] but lacking essential components to select for plasmids. Yeast cells were grown into mid-exponential phase (OD600 nm = 1.5) at 30 °C.