Protocol to quantify polyphosphate in human cell lines using a tagged PPBD peptide

Jiménez, Javier; Lázaro, Blanca; Sarrias, Ana; Tadeo, Francisco J.; Pérez-Montero, Marta; Clotet, Josep; Bru, Samuel

doi:10.1016/j.xpro.2022.101363

Cited by 2 publications

(3 citation statements)

References 3 publications

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“…However, binding of DAPI to inositol pyrophosphates causes similar shifts, which introduces a confounding variable [125]. Other groups have used the Xpress epitope tag fused to the polyP-binding domain (PPBD) of the E. coli PPX enzyme as a probe in fixed cells [126]. In a second step, the Xpress tag is detected with an antibody coupled to a fluorescent dye [126].…”

Section: Prunementioning

confidence: 99%

“…Other groups have used the Xpress epitope tag fused to the polyP-binding domain (PPBD) of the E. coli PPX enzyme as a probe in fixed cells [126]. In a second step, the Xpress tag is detected with an antibody coupled to a fluorescent dye [126]. Fluorescent polyPbinding molecules JC-D7 and JC-D8 have been developed for use in live cell imaging [127].…”

Section: Prunementioning

confidence: 99%

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The emerging landscape of eukaryotic polyphosphatases

McCarthy

Downey

2023

FEBS Letters

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Polyphosphate (polyP) is a conserved polymer of inorganic phosphate residues that can reach thousands of moieties in length. PolyP has been implicated in cellular functions ranging from energy and phosphate homeostasis to cell signalling in eukaryotes from yeast to humans. Despite the interest in the role of polyP as a signalling molecule, the spatiotemporal regulation of polyP itself remains poorly understood. This knowledge gap limits our ability to understand how polyP impacts the physiology of normal and diseased cells and how this might be exploited in a therapeutic context. Polyphosphatases, enzymes that degrade polyP to generate shorter chains and free inorganic phosphate are ideally positioned to mediate polyP dynamics. However, little is known about how the activities of these enzymes are linked to specific cellular functions and how they might be regulated. Here, we provide an indepth overview of polyphosphatase enzymes in budding yeast, which has served as a workhorse for polyP research, and in mammalian cells where the enzymes that make and degrade polyP have remained elusive. We identify critical open questions in both systems and propose strategies to guide future work.

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Section: Prunementioning

confidence: 99%

Section: Prunementioning

confidence: 99%

The emerging landscape of eukaryotic polyphosphatases

McCarthy

Downey

2023

FEBS Letters

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“…Inorganic polyphosphates (polyP) are polymers composed of orthophosphate residues connected by phosphoanhydride linkages like those found in ATP (Dedkova et al, 2016). PolyP has been detected in all species studied thus far, but its concentration and size vary greatly depending on the type of organism (Jiménez et al, 2022). PolyP's biological roles in unicellular prokaryotes and eukaryotes have been examined in more profundity.…”

Section: Introductionmentioning

confidence: 99%

Inorganic polyphosphate's role in energy production and mitochondrial permeability transition pore opening in tick mitochondria

Domingues¹,

Moraes

Fonseca

et al. 2023

Arch Insect Biochem Physiol

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Inorganic polyphosphate (polyP) is a biopolymer composed of phosphoanhydride‐linked orthophosphate molecules. PolyP is engaged in a variety of cellular functions, including mitochondrial metabolism. Here, we examined the effects of polyP on electron transport chain enzymes and F1Fo ATP synthase in tick embryos during embryonic development. The study found that polyPs containing medium and long chains (polyP15 and polyP65) enhanced the activity of complex I, complex II, complex III, and F1Fo ATP synthase, while short polyP chains (polyP3) had no effect. The study also examined the activity of exopolyphosphatases (PPX) in various energy‐demand situations. PPX activity was stimulated when ADP concentrations are high, characterizing a low‐energy context. When complexes I–III and F1Fo ATP synthase inhibitors were added in energized mitochondria, PPX activity decreased, whereas the mitochondrial uncoupler FCCP had no impact on PPX activity. Additionally, the study investigated the effect of polyP on mitochondrial swelling, finding that polyP causes mitochondrial swelling by increasing calcium effects on the mitochondrial permeability transition pore. The findings presented here to increase our understanding of the function of polyP in mitochondrial metabolism and its relationship to mitochondrial permeability transition pore opening in an arthropod model.

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Protocol to quantify polyphosphate in human cell lines using a tagged PPBD peptide

Cited by 2 publications

References 3 publications

The emerging landscape of eukaryotic polyphosphatases

The emerging landscape of eukaryotic polyphosphatases

Inorganic polyphosphate's role in energy production and mitochondrial permeability transition pore opening in tick mitochondria

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