scholarly journals A genetic screen for suppressors of hyper-repression of the fission yeast PHO regulon by Pol2 CTD mutation T4A implicates inositol 1-pyrophosphates as agonists of precocious lncRNA transcription termination

2020 ◽  
Vol 48 (19) ◽  
pp. 10739-10752
Author(s):  
Angad Garg ◽  
Stewart Shuman ◽  
Beate Schwer
Keyword(s):  
Fission Yeast ◽  
Rna Polymerase Ii ◽  
Genetic Screen ◽  
Pho Regulon ◽  
Wild Type ◽  
Phosphate Homeostasis ◽  
Inositol Pyrophosphate ◽  
Inositol 1 ◽  
Rna Polymerase Ii Ctd

Abstract Fission yeast phosphate homeostasis genes are repressed in phosphate-rich medium by transcription of upstream lncRNAs that interferes with activation of the flanking mRNA promoters. lncRNA control of PHO gene expression is influenced by the Thr4 phospho-site in the RNA polymerase II CTD and the 3′ processing/termination factors CPF and Rhn1, mutations of which result in hyper-repression of the PHO regulon. Here, we performed a forward genetic screen for mutations that de-repress Pho1 acid phosphatase expression in CTD-T4A cells. Sequencing of 18 independent STF (Suppressor of Threonine Four) isolates revealed, in every case, a mutation in the C-terminal pyrophosphatase domain of Asp1, a bifunctional inositol pyrophosphate (IPP) kinase/pyrophosphatase that interconverts 5-IP7 and 1,5-IP8. Focused characterization of two STF strains identified 51 coding genes coordinately upregulated vis-à-vis the parental T4A strain, including all three PHO regulon genes (pho1, pho84, tgp1). Whereas these STF alleles—asp1-386(Stop) and asp1-493(Stop)—were lethal in a wild-type CTD background, they were viable in combination with mutations in CPF and Rhn1, in which context Pho1 was also de-repressed. Our findings implicate Asp1 pyrophosphatase in constraining 1,5-IP8 or 1-IP7 synthesis by Asp1 kinase, without which 1-IPPs can accumulate to toxic levels that elicit precocious termination by CPF/Rhn1.

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

RNA ◽  
2015 ◽  
Vol 21 (10) ◽  
pp. 1770-1780 ◽  
Author(s):  
Beate Schwer ◽  
Ana M. Sanchez ◽  
Stewart Shuman
Keyword(s):  
Rna Polymerase ◽  
Fission Yeast ◽  
Rna Polymerase Ii ◽  
Phosphate Homeostasis ◽  
Rna Polymerase Ii Ctd

scholarly journals The INO2 and INO4 loci of Saccharomyces cerevisiae are pleiotropic regulatory genes.

1984 ◽  
Vol 4 (11) ◽  
pp. 2479-2485 ◽  
Author(s):  
B S Loewy ◽  
S A Henry
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Product ◽  
Regulatory Genes ◽  
Wild Type ◽  
Inositol 1 ◽  
Cytoplasmic Enzyme ◽  
Pleiotropic Phenotype ◽  
Reduced Rate ◽  
Phosphate Synthase

We isolated a mutant of Saccharomyces cerevisiae defective in the formation of phosphatidylcholine via methylation of phosphatidylethanolamine. The mutant synthesized phosphatidylcholine at a reduced rate and accumulated increased amounts of methylated phospholipid intermediates. It was also found to be auxotrophic for inositol and allelic to an existing series of ino4 mutants. The ino2 and ino4 mutants, originally isolated on the basis of an inositol requirement, are unable to derepress the cytoplasmic enzyme inositol-1-phosphate synthase (myo-inositol-1-phosphate synthase; EC 5.5.1.4). The INO4 and INO2 genes were, thus, previously identified as regulatory genes whose wild-type product is required for expression of the INO1 gene product inositol-1-phosphate synthase (T. Donahue and S. Henry, J. Biol. Chem. 256:7077-7085, 1981). In addition to the identification of a new ino4-allele, further characterization of the existing series of ino4 and ino2 mutants, reported here, demonstrated that they all have a reduced capacity to convert phosphatidylethanolamine to phosphatidylcholine. The pleiotropic phenotype of the ino2 and ino4 mutants described in this paper suggests that the INO2 and INO4 loci are involved in the regulation of phospholipid methylation in the membrane as well as inositol biosynthesis in the cytoplasm.

scholarly journals RNA polymerase II CTD interactome with 3′ processing and termination factors in fission yeast and its impact on phosphate homeostasis

2018 ◽  
Vol 115 (45) ◽  
pp. E10652-E10661 ◽  
Author(s):  
Ana M. Sanchez ◽  
Stewart Shuman ◽  
Beate Schwer
Keyword(s):  
Gene Expression ◽  
Rna Polymerase ◽  
Fission Yeast ◽  
Rna Polymerase Ii ◽  
Noncoding Rna ◽  
Synthetically Lethal ◽  
Heptad Repeats ◽  
Carboxy Terminal ◽  
Key Steps ◽  
Rna Polymerase Ii Ctd

The carboxy-terminal domain (CTD) code encrypted within the Y1S2P3T4S5P6S7heptad repeats of RNA polymerase II (Pol2) is deeply rooted in eukaryal biology. Key steps to deciphering the code are identifying the events in gene expression that are governed by individual “letters” and then defining a vocabulary of multiletter “words” and their meaning. Thr4 and Ser7 exert opposite effects on the fission yeastpho1gene, expression of which is repressed under phosphate-replete conditions by transcription of an upstream flanking long noncoding RNA (lncRNA). Here we attribute the derepression ofpho1by a CTD-S7Amutation to precocious termination of lncRNA synthesis, an effect that is erased by mutations of cleavage-polyadenylation factor (CPF) subunits Ctf1, Ssu72, Ppn1, Swd22, and Dis2 and termination factor Rhn1. By contrast, a CTD-T4Amutation hyperrepressespho1, as do CPF subunit and Rhn1 mutations, implying thatT4Areduces lncRNA termination. Moreover, CTD-T4Ais synthetically lethal withppn1∆ andswd22∆, signifying that Thr4 and the Ppn1•Swd22 module play important, functionally redundant roles in promoting Pol2 termination. We find that Ppn1 and Swd22 become essential for viability when the CTD array is curtailed and thatS7Aovercomes the need for Ppn1•Swd22 in the short CTD context. Mutational synergies highlight redundant essential functions of (i) Ppn1•Swd22 and Rhn1, (ii) Ppn1•Swd22 and Ctf1, and (iii) Ssu72 and Dis2 phosphatases. CTD allelesY1F,S2A, andT4Ahave overlapping synthetic lethalities withppn1∆ andswd22∆, suggesting that Tyr1-Ser2-Thr4 form a three-letter CTD word that abets termination, with Rhn1 being a likely “reader” of this word.

scholarly journals Genetic and structural analysis of the essential fission yeast RNA polymerase II CTD phosphatase Fcp1

RNA ◽  
2015 ◽  
Vol 21 (6) ◽  
pp. 1135-1146 ◽  
Author(s):  
Beate Schwer ◽  
Agnidipta Ghosh ◽  
Ana M. Sanchez ◽  
Christopher D. Lima ◽  
Stewart Shuman
Keyword(s):  
Structural Analysis ◽  
Rna Polymerase ◽  
Fission Yeast ◽  
Rna Polymerase Ii ◽  
Ctd Phosphatase ◽  
Rna Polymerase Ii Ctd

Interaction between the Cig1 and Cig2 B-type cyclins in the fission yeast cell cycle

1994 ◽  
Vol 14 (1) ◽  
pp. 768-776
Author(s):  
T Connolly ◽  
D Beach
Keyword(s):  
Cell Cycle ◽  
Fission Yeast ◽  
Null Allele ◽  
Wild Type Strain ◽  
Periodic Oscillation ◽  
S Phase ◽  
Wild Type ◽  
Nuclear Separation ◽  
The Relationship

In this report, we describe the cloning and characterization of a B-type cyclin, Cig2 from the fission yeast Schizosaccharomyces pombe. The cig2 gene encodes a 45-kDa protein that is most similar to a previously identified B-type cyclin in S. pombe, Cdc13. Deletion of cig2 had no observable effect on cell viability or progression through the cell cycle. Strains carrying the cig2 null allele do, however, exhibit an enhanced ability to undergo conjugation relative to a wild-type strain. The cig2 transcript was found to undergo periodic oscillation during the cell cycle, peaking at the G1/S-phase boundary. We have investigated the relationship between Cig2 and the other B-type cyclins, Cig1 and Cdc13, in the fission yeast. We found that cells carrying disruptions of both the cig1 and cig2 genes contain multiple nuclei with a 1C DNA content, suggesting that they are delayed in progression through the G1 phase of the cell cycle. The phenotype of this double mutant suggests that there is a delay in septum formation, possibly as a result of defective nuclear separation.

The INO2 and INO4 loci of Saccharomyces cerevisiae are pleiotropic regulatory genes

1984 ◽  
Vol 4 (11) ◽  
pp. 2479-2485
Author(s):  
B S Loewy ◽  
S A Henry
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Product ◽  
Regulatory Genes ◽  
Wild Type ◽  
Inositol 1 ◽  
Cytoplasmic Enzyme ◽  
Pleiotropic Phenotype ◽  
Reduced Rate ◽  
Phosphate Synthase

We isolated a mutant of Saccharomyces cerevisiae defective in the formation of phosphatidylcholine via methylation of phosphatidylethanolamine. The mutant synthesized phosphatidylcholine at a reduced rate and accumulated increased amounts of methylated phospholipid intermediates. It was also found to be auxotrophic for inositol and allelic to an existing series of ino4 mutants. The ino2 and ino4 mutants, originally isolated on the basis of an inositol requirement, are unable to derepress the cytoplasmic enzyme inositol-1-phosphate synthase (myo-inositol-1-phosphate synthase; EC 5.5.1.4). The INO4 and INO2 genes were, thus, previously identified as regulatory genes whose wild-type product is required for expression of the INO1 gene product inositol-1-phosphate synthase (T. Donahue and S. Henry, J. Biol. Chem. 256:7077-7085, 1981). In addition to the identification of a new ino4-allele, further characterization of the existing series of ino4 and ino2 mutants, reported here, demonstrated that they all have a reduced capacity to convert phosphatidylethanolamine to phosphatidylcholine. The pleiotropic phenotype of the ino2 and ino4 mutants described in this paper suggests that the INO2 and INO4 loci are involved in the regulation of phospholipid methylation in the membrane as well as inositol biosynthesis in the cytoplasm.

scholarly journals Interaction between the Cig1 and Cig2 B-type cyclins in the fission yeast cell cycle.

1994 ◽  
Vol 14 (1) ◽  
pp. 768-776 ◽  
Author(s):  
T Connolly ◽  
D Beach
Keyword(s):  
Cell Cycle ◽  
Fission Yeast ◽  
Null Allele ◽  
Wild Type Strain ◽  
Periodic Oscillation ◽  
S Phase ◽  
Wild Type ◽  
Nuclear Separation ◽  
The Relationship

In this report, we describe the cloning and characterization of a B-type cyclin, Cig2 from the fission yeast Schizosaccharomyces pombe. The cig2 gene encodes a 45-kDa protein that is most similar to a previously identified B-type cyclin in S. pombe, Cdc13. Deletion of cig2 had no observable effect on cell viability or progression through the cell cycle. Strains carrying the cig2 null allele do, however, exhibit an enhanced ability to undergo conjugation relative to a wild-type strain. The cig2 transcript was found to undergo periodic oscillation during the cell cycle, peaking at the G1/S-phase boundary. We have investigated the relationship between Cig2 and the other B-type cyclins, Cig1 and Cdc13, in the fission yeast. We found that cells carrying disruptions of both the cig1 and cig2 genes contain multiple nuclei with a 1C DNA content, suggesting that they are delayed in progression through the G1 phase of the cell cycle. The phenotype of this double mutant suggests that there is a delay in septum formation, possibly as a result of defective nuclear separation.

scholarly journals Distinct requirement of RNA polymerase II CTD phosphorylations in budding and fission yeast

Transcription ◽  
2012 ◽  
Vol 3 (5) ◽  
pp. 231-234 ◽  
Author(s):  
Clément Cassart ◽  
Julie Drogat ◽  
Valérie Migeot ◽  
Damien Hermand
Keyword(s):  
Rna Polymerase ◽  
Fission Yeast ◽  
Rna Polymerase Ii ◽  
Rna Polymerase Ii Ctd
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