Azevedo, C;
Desfougères, Y;
Jiramongkol, Y;
Partington, H;
Trakansuebkul, S;
Singh, J;
Steck, N;
... Saiardi, A; + view all
(2019)
Development of a yeast model to study the contribution of vacuolar polyphosphate metabolism to lysine polyphosphorylation.
Journal of Biological Chemistry
10.1074/jbc.RA119.011680.
(In press).
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Abstract
A recently discovered protein post-translational modification, lysine polyphosphorylation (K-PPn), consists of the covalent attachment of inorganic polyphosphate (polyP) to lysine residues. The non-enzymatic nature of K-PPn means that the degree of this modification depends on both polyP abundance and the amino acids surrounding the modified lysine. K-PPn was originally discovered in budding yeast (Saccharomyces cerevisiae), in which polyP anabolism and catabolism are well characterized. However, yeast vacuoles accumulate large amounts of polyP, and upon cell lysis, the release of the vacuolar polyP could non-physiologically cause K-PPn of nuclear and cytosolic targets. Moreover, yeast vacuoles possess two very active endopolyphosphatases, Ppn1 and Ppn2, that could have opposing effects on the extent of K-PPn. Here, we characterized the contribution of vacuolar polyP metabolism to K-PPn of two yeast proteins, Top1 (DNA topoisomerase 1) and Nsr1 (nuclear signal recognition 1). We discovered that whereas Top1-targeting K-PPn is only marginally affected by vacuolar polyP metabolism, Nsr1-targeting K-PPn is highly sensitive to the release of polyP and of endopolyphosphatases from the vacuole. Therefore, to better study K-PPn of cytosolic and nuclear targets, we constructed a yeast strain devoid of vacuolar polyP by targeting the exopolyphosphatase Ppx1 to the vacuole and concomitantly depleting the two endopolyphosphatases (ppn1Δppn2Δ, vt-Ppx1). This strain enabled us to study K-PPn of cytosolic and nuclear targets without the interfering effects of cell lysis on vacuole polyP and of endopolyphosphatases. Furthermore, we also define the fundamental nature of the acidic amino acid residues to the K-PPn target domain.
| Type: | Article |
|---|---|
| Title: | Development of a yeast model to study the contribution of vacuolar polyphosphate metabolism to lysine polyphosphorylation |
| Location: | United States |
| Open access status: | An open access version is available from UCL Discovery |
| DOI: | 10.1074/jbc.RA119.011680 |
| Publisher version: | https://doi.org/10.1074/jbc.RA119.011680 |
| Language: | English |
| Additional information: | This version is the author accepted manuscript. For information on re-use, please refer to the publisher’s terms and conditions. |
| Keywords: | Inorganic polyphosphate, Lysine modifications, Top1, cell signaling, lysine polyphosphorylation (K-PPn), molecular cell biology, phosphorylation, polyphosphatases, post-translational modification (PTM), protein phosphorylation |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Lab for Molecular Cell Bio MRC-UCL UCL > Provost and Vice Provost Offices > UCL BEAMS UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Maths and Physical Sciences UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Maths and Physical Sciences > Dept of Chemistry |
| URI: | https://discovery.ucl.ac.uk/id/eprint/10088335 |
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