# An Update on Polyphosphate In Vivo Activities
## 6 Mammalians  
Mammalian cells contain polyP, although the cellular content of the polymer, its subcellular localisation, and chain length greatly vary [1]. Using [32P]-radiolabeled polymers has shown that polyP content in rat hepatocytes is highest in the nucleus (89 \(\pm\) 7 uM) followed by plasma membranes (43 \(\pm\) 3 uM), cytosol (12 \(\pm\) 2 uM), mitochondria (11 \(\pm\) 0.6 uM) and lowest in microsomes (4 \(\pm\) 0.5 uM) [5]. High polyP levels in the cytosol have also been found in various immortalised cultured cell lines, including NIH3T3 fibroblasts, Vero epithelial kidney, and Jurkat CD4+ T cells [5]. In contrast to simpler organisms where polyP metabolising enzymes have been well characterised, the polyP synthesising and degrading machinery in mammalians have remained enigmatic. Surprisingly, and despite the fundamental functions of the polymer, the polyp-metabolising enzymes of simpler organisms are not conserved in mammalians, and knowledge about enzymes involved in polyP turnover is just beginning to emerge [79]. Mammalian phospholipase D shares similarities with bacterial PPK1 and PPK2 but has no polyP-modifying activity [7]. Bacterial PPX1 and human (h)-Prune are functionally equivalent but non-homologous proteins originating from distinct ancestors. PPX1 is a member of the PPX-GppA phosphatases family. At the same time, h-Prune belongs to the DHH phosphoesterases superfamily, which shares the phosphodiesterase activity to hydrolyses short-chain polyP (up to 4 P\({}_{\text{i}}\)). In contrast to PPX, h-Prune processes long-chain molecules with >25 P\({}_{\text{i}}\) residues poorly. Consistently, recombinant h-Prune is unable to hydrolyse short- and medium-chain polyP. Conversely, the knockdown of h-Prune expression significantly decreases polyP and ATP content in treated HEK293 cells [52]. Prune is involved in mammalian polyP metabolism, although its exact role and regulation have not been fully elucidated [52; 80]. Prune is expressed and functions in polyP metabolism in _Drosophila melanogaster_, suggesting that some fundamental principles of polyP regulation are conserved throughout evolution. However, the cellular localisation of Prune differs among the species. In _Drosophila melanogaster_, Prune is predominantly in mitochondria. In contrast, in addition to mitochondria, the polymer is enriched in various other compartments, including ER, cytoplasm, nucleus, and many (secretory) granules and vesicles in mammalian cells [80]. Throughout the species, polyP is enriched in mitochondria and couples phosphate- with ATP-homeostasis. In mammalians, the FOF1-ATP synthase, which generates ATP within the mitochondrial matrix, modulates polyP content and chain length [80; 81]. Another group of conserved polyp-metabolising enzymes are alkaline phosphatases (Table 1), ubiquitous membrane-bound glycoproteins that are capable of cleaving phosphoester bonds and additionally have the capacity for degrading polyP [82]. Shrimp alkaline phosphatase efficiently degrades extracellular polyP in plasma, confirming that the enzyme has polyp-metabolising activities [60]. Alkaline phosphatases are similar to yeast cytosolic and nuclear diphosphoitol polyphosphate phosphohydrolase 1 and 2 (DDP1 and DDP2) [83]. In addition to polyP synthesis and degradation, the polymer is also regulated by the availability of P\({}_{\text{i}}\). The Xenotropic and polytropic retrovirus receptor 1 (XPR1) was initially identified as a docking receptor for retroviruses. However, later studies showed that XPR1 also exports P\({}_{\text{i}}\) in human and murine platelets [84; 85]. XPR1 shares homology with Pho84, Pho87, Pho90, and Pho89 P\({}_{\text{i}}\) transporters in _Saccharomyces cerevisiae_[86]. XPR1 is an atypical G-protein-coupled receptor expressed in intracellular organelles and at the plasma membrane. In addition to being regulated by XPR1, polyP itself modulates membrane channels. Primary rat hepatocytes express the mitochondria permeability transition pore (mPTP) at their inner mitochondrial membrane.