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Cross-regulation of Iron and Phosphate Homeostasis by Fep1 and Pho7 in Fission yeast [RNA-seq]

ABSTRACT: Iron and phosphate are essential micronutrients with partially interconnected homeostatic control; however, the underlying transcriptional mechanisms that integrate these signals in the fission yeast Schizosaccharomyces pombe, remain unclear. Fep1 (GATA-type repressor) is a key regulator of iron-uptake genes, whereas Pho7 (Zn2Cys6-family transcription factor) drives phosphate starvation response. However, whether each of two factors influences the target regulon controlled by the other remains unclear. We combined genome-wide chromatin immunoprecipitation sequencing, RNA sequencing, and auxin-inducible degron-mediated acute depletion to define the joint Fep1–Pho7 transcriptional control and regulatory landscapes under defined iron and phosphate conditions. Unexpectedly, these analyses showed extensive overlap in chromatin occupancy between Fep1 and Pho7 at a subset of target promoters. Fep1 bound not only to canonical iron-uptake genes but also the pho7+ locus and multiple phosphate-responsive promoters, whereas Pho7 showed association with a defined subset of Fep1 targets. Acute depletion experiments showed that Fep1 loss substantially increased Pho7 chromatin occupancy at iron-responsive promoters. Under iron-depleted conditions, Pho7 contributed to the transcriptional activation of specific Fep1 target genes (fio1+/fip1+ and str3+), which indicated that Pho7 supports iron uptake gene expression when Fep1 repression is attenuated. Condition-dependent binding analyses showed that iron availability influences the relative regulatory contribution of the two factors at shared loci. These findings support a model in which Fep1 and Pho7 form a cross-regulatory module that converges on shared genes to enable iron-dependent redistribution of their influence. Together, these findings suggest a mechanistic framework for integrating iron and phosphate signals at the promoter level in fission yeast.

ORGANISM(S): Schizosaccharomyces pombe

PROVIDER: GSE344406 | GEO | 2026/08/20

REPOSITORIES: GEO

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Project description:Inorganic phosphate is an essential nutrient required by organisms for growth. During phosphate starvation, Saccharomyces cerevisiae activates the phosphate signal transduction (PHO) pathway leading to the expression of the secreted acid phosphatase, PHO5. The fission yeast, Schizosaccharomyces pombe, regulates expression of the ScPHO5 homolog (pho1+) via a non-orthologous PHO pathway. The genes induced by phosphate limitation and the molecular mechanism by which the genetically identified positive (pho7+) and negative (csk1+) regulators function are not known. Here we use a combination of molecular biology, expression microarrays, and chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-Seq) to characterize the role of pho7+ and csk1+ in the PHO response. We define the set of genes that comprise the initial response to phosphate starvation in S. pombe. We identify a conserved PHO response for the ScPHO5 (pho1+), ScPHO84 (spbc8e4.01c+), and ScGIT1 (spbc1271.09+) orthologs. We use ChIP-Seq to identify members of the Pho7 regulon and characterize Pho7 binding in response to phosphate-limitation and Csk1 activity. We demonstrate that activation of pho1+ requires Pho7 binding to a UAS in the pho1+ promoter and that Csk1 repression does not regulate Pho7 enrichment. Further, we find that Pho7-dependent activation is not limited to phosphate-starvation, as additional environmental stress response pathways require pho7+ for maximal induction. We provide a global analysis of the PHO pathway in S. pombe. Our results elucidate the conserved core regulon required for responding to phosphate starvation between distantly related ascomycetes and a better understanding of flexibility in environmental stress response networks. Schizosaccharomyces pombe 972 h- cells were starved for inorganic phosphate for 0, 30, 60, 120, or 240 minutes pior to microarray preparation to determine the extent and temporal resolution of the phosphate starvation response. At 120 minutes post-starvation we define a set of genes that are directly and specifically induced by phosphate starvation, providing a time-point at which all other experiments were performed. We characterize the pho7+- and csk1+- dependency of this PHO response at 120 minutes post-starvation in pho7+csk1+, pho7M-NM-^T, csk1M-NM-^T, and pho7M-NM-^Tcsk1M-NM-^T cells. In a seperate set of experiments we characterized the S. pombe stress response to copper limitation, iron limitation, and carbon switching at 120 minutes post-stress and osmotic shift at 20 minutes post-stress in both pho7+ and pho7M-NM-^T cells.

Project description:The primary structure and phosphorylation pattern of the tandem YSPTSPS repeats of the RNA polymerase II CTD comprise an informational code that coordinates transcription, chromatin modification, and RNA processing. To gauge the contributions of individual CTD coding M-bM-^@M-^\lettersM-bM-^@M-^] to gene expression, we analyzed the poly(A)+ transcriptomes of fission yeast mutants that lack each of the four inessential CTD phosphoacceptors: Tyr1, Ser2, Thr4, and Ser7. There was a hierarchy of CTD mutational effects with respect to the number of dysregulated protein-coding RNAs, with S2A (n=227) >> Y1F (n=71) > S7A (n=58) >> T4A (n=7). The majority of the protein-coding RNAs affected in Y1F cells were coordinately affected by S2A, suggesting that Tyr1-Ser2 constitutes a two-letter code M-bM-^@M-^\wordM-bM-^@M-^]. Y1F and S2A elicited increased expression of genes encoding proteins involved in iron uptake (Frp1, Fip1, Fio1, Str3, Str1, Sib1), without affecting the expression of the genes that repress the iron regulon, implying that Tyr1-Ser2 transduces a repressive signal. Y1F and S2A cells had increased levels of ferric reductase activity and were hypersensitive to phleomycin, indicative of elevated intracellular iron. The T4A and S7A mutations had opposing effects on the phosphate response pathway. T4A reduced the expression of two genes encoding proteins involved in phosphate acquisition (the Pho1 acid phosphatase and the phosphate transporter SPBC8E4.01c), without affecting the expression of known genes that regulate the phosphate response pathway, while S7A increased pho1+ expression. Meiotic genes were enriched among those up-regulated in S7A cells. These results highlight specific cellular gene expression programs that are responsive to distinct CTD cues. Interrogation of the S. pombe transcriptome using polyA+ strand specific RNA sequencing (Illumina HiSeq 2000) in cultures. A total of 16 samples were analysed: two biological repeates of each WT, Y1F, S2A, T4A, S7A,Y1F-S7A, S2A-S7A and T4A-S7A strains

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Cross-regulation of Iron and Phosphate Homeostasis by Fep1 and Pho7 in Fission yeast [RNA-seq]

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