Inositol pyrophosphates impact phosphate homeostasis via modulation of RNA 3’ processing and transcription termination

Abstract

Fission yeast phosphate acquisition genes pho1, pho84, and tgp1 are repressed in phosphate-rich medium by transcription of upstream lncRNAs. Here we show that phosphate homeostasis is subject to metabolite control by inositol pyrophosphates (IPPs), exerted through the 3'-processing/termination machinery and the Pol2 CTD code. Increasing IP8 (via Asp1 IPP pyrophosphatase mutation) de-represses the PHO regulon and leads to precocious termination of prt lncRNA synthesis. pho1 de-repression by IP8 depends on cleavage-polyadenylation factor (CPF) subunits, termination factor Rhn1, and the Thr4 letter of the CTD code. pho1 de-repression by mutation of the Ser7 CTD letter depends on IP8. Simultaneous inactivation of the Asp1 and Aps1 IPP pyrophosphatases is lethal, but this lethality is suppressed by mutations of CPF subunits Ppn1, Swd22, Ssu72, and Ctf1 and CTD mutation T4A. Failure to synthesize IP8 (via Asp1 IPP kinase mutation) results in pho1 hyper-repression. Synthetic lethality of asp1∆ with Ppn1, Swd22, and Ssu72 mutations argues that IP8 plays an important role in essential 3'processing/termination events, albeit in a manner genetically redundant to CPF. Transcriptional profiling delineates an IPP-responsive regulon composed of genes overexpressed when IP8 levels are increased. Our results establish a novel role for IPPs in cell physiology.

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

“…Transcriptional profiling uncovered an ensemble of 51 coding genes that were coordinately up-regulated in two different STF nonsense mutant strains; this set includes all three genes of the PHO regulon. Thus, our forward genetic screen fortifies the recent proposal ( 29 ) that fission yeast phosphate homeostasis is subject to metabolite control by inositol pyrophosphates, exerted via the 3′ processing/termination machinery and the Pol2 CTD.…”

“…Recent studies show that failure to synthesize IP8 and 1-IP7 in Asp1 null or Asp1 kinase-defective strains results in hyper-repression of the PHO regulon under phosphate-replete conditions ( 29 ). Synthetic lethalities of asp1 Δ and asp1 kinase-defective alleles with mutations of CPF subunits Ppn1, Swd22, and Ssu72 point to an important role for IP8 (or 1-IP7) in essential 3′ processing/termination events, albeit in a manner genetically redundant to CPF ( 29 ). Pertinent to the present suppressor analysis is the observation that asp1 Δ and the kinase-defective allele are synthetically lethal with CTD-T4A (B. Schwer, unpublished), i.e.…”

“…Two other genes involved in phosphate acquisition were also upregulated: SPBPB2B2.06c encoding an extracellular 5′ nucleotidase (up 28- to 48-fold) and SPBC1683.01 encoding a transmembrane phosphate transporter (up 3- to 4-fold) (Figure 7 ). It is suggested that ecl3 and SPBC1683.01, in common with the three PHO genes, might be subject to transcription interference by 5′ flanking lncRNAs ( 29 ).…”

Conserved protein Pir2ARS2 mediates gene repression through cryptic introns in lncRNAs

“…Transcription termination and degradation of the lncRNAs prevents them from invading and repressing downstream genes 7,[11][12][13][14] . However, under specific growth conditions, readthrough transcription of lncRNAs leads to repression of downstream genes 15 . Underscoring a direct role, cells defective in lncRNA production show de-repression of target genes [6][7][8]11,12 .…”

Rapid stimulation of cellular Pi uptake by the inositol pyrophosphate InsP8induced by its photothermal release from lipid nanocarriers using a near infra-red light-emitting diode

“…In the absence of specic inhibitors of PPIP5K-mediated 1,5-InsP 8 synthesis, most previous advances into the biological activities of this particular PP-InsP have primarily been driven by long-term genetic perturbations of PPIP5K expression. 11,31,[57][58][59][60] In contrast, we have described a method that can illuminate molecular actions of 1,5-InsP 8 within a time-frame of a few minutes. Our methodology uses near infra-red irradiation which is more compatible with biological samples than is UV irradiation in uncaging experiments, which can generate damaging radicals.…”