Systematic Screen of Schizosaccharomyces pombe Deletion Collection Uncovers Parallel Evolution of the Phosphate Signal Transduction Pathway in Yeasts

Abstract

The phosphate signal transduction (PHO) pathway, which regulates genes in response to phosphate starvation, is well defined in Saccharomyces cerevisiae. We asked whether the PHO pathway was the same in the distantly related fission yeast Schizosaccharomyces pombe. We screened a deletion collection for mutants aberrant in phosphatase activity, which is primarily a consequence of pho1 ؉ transcription. We identified a novel zinc finger-containing protein (encoded by spbc27b12.11c ؉ ), which we have named pho7 ؉ , that is essential for pho1؉ transcriptional induction during phosphate starvation. Few of the S. cerevisiae genes involved in the PHO pathway appear to be involved in the regulation of the phosphate starvation response in S. pombe. Only the most upstream genes in the PHO pathway in S. cerevisiae (ADO1, DDP1, and PPN1) share a similar role in both yeasts. Because ADO1 and DDP1 regulate ATP and IP 7 levels, we hypothesize that the ancestor of these yeasts must have sensed similar metabolites in response to phosphate starvation but have evolved distinct mechanisms in parallel to sense these metabolites and induce phosphate starvation genes.The cellular homeostasis of inorganic phosphate is required for optimal growth and efficient metabolism. The response of the model organism Saccharomyces cerevisiae to extracellular phosphate starvation is well characterized and mediated by the phosphate signal transduction (PHO) pathway (20,24). To determine whether the PHO pathway is conserved in other Ascomycota fungal species, we screened for PHO pathway mutants in the evolutionarily distantly related Schizosaccharomyces pombe, which last shared a common ancestor with S. cerevisiae more than 1 billion years ago (7).The PHO pathway in S. cerevisiae often is defined by the regulation of PHO5, which encodes a phosphate starvationregulated acid phosphatase (17,20). PHO5 is highly induced during phosphate starvation. ScPho5 activity is detected using a diazo-coupling assay with 1-napthylphosphate (9). Numerous studies have determined that PHO5 transcription is regulated by the specific transcription factors Pho4 and Pho2 and by more general chromatin remodeling complexes, such as SWI/ SNF, SAGA, and INO80 (1, 16, 31). Pho4 localization and activity is regulated by a cyclin/cyclin-dependent kinase complex (Pho81/Pho80/Pho85) (11,12,25). During high extracellular phosphate conditions, the kinase complex is active and phosphorylates Pho4, leading to nuclear exclusion and little transcription of PHO5 (10, 15). During low extracellular phosphate conditions, Pho81 inhibits the kinase complex through a noncovalent interaction with IP 7 (inositol heptakisphosphate) (18,19). Certain isomers of IP 7 increase in abundance in response to phosphate starvation, although how extracellular phosphate concentration leads to these increases is unclear. However, Vip1 is required to phosphorylate IP 6 to form 4-PP-IP 5 or 6-PP-IP 5 , and Ddp1 is required for dephosphorylation back to IP 6 (19). Increases in IP 7 during extracellular phosphate st...

RNA polymerase II CTD phospho-sites Ser5 and Ser7 govern phosphate homeostasis in fission yeast

“…6). In contrast, csk1Δ cells constitutively express acid phosphatase in phosphate-replete conditions (Henry et al 2011;Carter-O'Connell et al 2012), at a level eightfold greater than wild type (Fig. 6).…”

“…Pho1 expression is strictly dependent on the yeast transcription factor Pho7 (Henry et al 2011;Carter-O'Connell et al 2012), such that pho7Δ cells have virtually no acid phosphatase activity, even when starved for phosphate (Fig. 6).…”

“…Upregulation of the expression of these genes during phosphate starvation depends on the transcription factor Pho7 (Henry et al 2011;Carter-O'Connell et al 2012), a member of the zinc binuclear cluster family of fungal DNA-binding transcription regulators (MacPherson et al 2006). The protein kinase Csk1 is a negative regulator of the phosphate response, as judged by the fact that pho1 expression is constitutively turned on in csk1Δ cells under phosphate-replete conditions (Henry et al 2011;Carter-O'Connell et al 2012;Schwer et al 2014). Csk1 is a CDK-activating kinase with several physiological targets, including the kinases Cdc2, Mcs6, and Cdk9 (Saiz and Fisher 2002;Pei et al 2006).…”

Individual letters of the RNA polymerase II CTD code govern distinct gene expression programs in fission yeast

“…The protein kinase Csk1 is a negative regulator of Pho1 and phosphate transporter expression, insofar as both are constitutively induced in csk1Δ cells. The fact that constitutive expression of Pho1 is abolished in a csk1Δ pho7Δ double mutant implies that Pho7 is essential for Pho1 transcription and acts downstream of Csk1 (17,18). The RNAseq data indicate that the decrements in Pho1 and phosphate transporter expression in T4A and T4A•S7A cells are not a consequence of decreased expression of the positive regulator Pho7 or increased expression of the negative regulator Csk1, because pho1 and csk1 mRNA levels were not affected significantly by the CTD mutations (Fig.…”

“…Pho1 and phosphate transporter expression are induced in response to phosphate starvation via the action of the Zn-finger transcription factor Pho7 (17,18). The protein kinase Csk1 is a negative regulator of Pho1 and phosphate transporter expression, insofar as both are constitutively induced in csk1Δ cells.…”