Genetic Screening for Regulators of Prz1, a Transcriptional Factor Acting Downstream of Calcineurin in Fission Yeast

• Published in: The Journal of biological chemistry 2012-06, Vol.287 (23), p.19294-19303
• Main Authors: Koike, Atsushi, Kato, Toshiaki, Sugiura, Reiko, Ma, Yan, Tabata, Yuki, Ohmoto, Koji, Sio, Susie O., Kuno, Takayoshi
• Format: Article
• Language: English
• Subjects: Calcineurin; Calcineurin - genetics; Calcineurin - metabolism; Cell Biology; Fungal Proteins - metabolism; Gene Expression Regulation, Fungal - physiology; Protein-Protein Interactions; Prz1; Schizosaccharomyces - genetics; Schizosaccharomyces - metabolism; Signal Transduction - physiology; Transcription Factors; Transcription Factors - genetics; Transcription Factors - metabolism; Transcription, Genetic - physiology; Yeast; Zinc Finger; Zinc Fingers
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M111.310615

Calcineurin phosphatase plays crucial roles in a wide variety of cell types and organisms. Dephosphorylation of the nuclear factor of activated T-cell (NFAT) family of transcriptional factors by calcineurin is essential for activating immune-responsive genes in mammals. NFAT activity is also regulated by diverse signaling pathways, which affect NFAT kinases and nuclear partner proteins. In fission yeast, calcineurin dephosphorylates and activates Prz1, a C2H2-type zinc finger transcriptional factor. Calcineurin-Prz1 signaling regulates the expression of the Pmc1 Ca2+ pump. Prz1-overexpressing cells showed extremely slow growth and high transcriptional activity of Prz1 in the absence of stimulation. Here, we isolated seven genes as dosage-dependent suppressors of this slow growth phenotype. These seven genes encode Rad24, Rad25, Pka1, Msn5 (SPAC328.01c), Pac1, Ape2, and Tfs1. All of them decreased the high transcriptional activity caused by Prz1 overexpression. Overexpression of Pka1, Rad24, and Rad25 also repressed the Ca2+-induced transcriptional activity in cells with Prz1 expressed at wild-type levels. Knock-out of rad24 or rad25 significantly enhanced the transcriptional activity of Prz1, whereas knock-out or mutation of other genes did not enhance the activity. The 14-3-3 proteins, Rad24 and Rad25, bound Prz1 and the Rad24-binding site located at residues 421–426 of Prz1. In msn5 deletion mutants, GFP-Prz1 localized at nucleus in the absence of Ca2+ stimulation, suggesting that Msn5 functions as an exportin for Prz1. In summary, our data suggest that Rad24 and Rad25 negatively regulate Prz1 and that Pka1, Msn5, Pac1, Tfs1, and Ape2 also regulate Prz1. The transcriptional factor Prz1 is activated by calcineurin, whereas other regulators of Prz1 are unknown. Pka1, Rad24, Rad25, Msn5, Pac1, Ape2, and Tfs1 are isolated as dosage-dependent suppressors of the slow growth phenotype caused by Prz1 overexpression. Prz1 is regulated by proteins involved in various biological processes. Identification of regulators of Prz1 is crucial for understanding calcineurin-mediated signal transduction.