Disease relevance of PNPT1

• Overexpression of hPNPase(OLD-35) in human melanoma cells and melanocytes induces distinctive changes associated with senescence, potentially mediated by direct degradation of c-myc mRNA by this enzyme. hPNPase(OLD-35) contains two RNase PH (RPH) domains, one PNPase domain, and two RNA binding domains [1].
• These results suggest that there is no enzyme similar to E. coli PAP I in spinach chloroplasts and that polyadenylation and exonucleolytic degradation of RNA in spinach chloroplasts are performed by one enzyme, PNPase [2].
• Our findings suggest that PNPase plays multifaceted roles in enhancing Yersinia survival in response to stressful conditions [3].
• Interferon-alpha (IFN-alpha) enhances the activity of the 5-fluorouracil (5-FU) prodrug 5'-deoxy-5-fluorouridine (5'-dFUrd) in colorectal cancer cells in vitro by upregulating the enzyme pyrimidine nucleoside phosphorylase (PNPase), which is responsible for converting 5'-dFUrd to 5-FU [4].
• Pnpt, the natural promoter driving expression of the kanamycin-resistance determinant from Tn5, was poorly expressed in Anabaena [5].

High impact information on PNPT1

• RNase E and PNPase are found in a multienzyme complex called the degradosome [6].
• Identification and cloning of human polynucleotide phosphorylase, hPNPase old-35, in the context of terminal differentiation and cellular senescence [7].
• We therefore conducted a mechanistic study of PNPase biogenesis in the mitochondrion [8].
• Mammalian PNPase exhibits exoribonuclease and poly(A) polymerase activities, and PNPase overexpression inhibits cell growth, induces apoptosis, and stimulates proinflammatory cytokine production [9].
• Combined, the data demonstrate an unexpected IMS localization and a key role for PNPase in maintaining mitochondrial homeostasis [9].

Biological context of PNPT1

• The activity of PNPase was directed to RNA degradation or polymerization by manipulating physiologically relevant concentrations of P(i) and ADP [2].
• Prior overexpression studies resulted in PNPase localization to both the cytosol and mitochondria, concurrent with cytosolic RNA degradation and pleiotropic cellular effects, including growth inhibition and apoptosis, that may not reflect a physiologic role for endogenous PNPase [8].
• Consistent with its IMS localization, PNPase knockdown with RNA interference did not affect mitochondrial RNA levels [9].
• However, PNPase reduction impaired mitochondrial electrochemical membrane potential, decreased respiratory chain activity, and was correlated with altered mitochondrial morphology [9].
• Genetic and molecular analysis of one such mutant revealed that it contains a mutation in the pnp gene, which encodes the PNPase catalytic subunit alpha [10].

Anatomical context of PNPT1

• Here, we demonstrate that the hPNPase is localized in mitochondria [11].
• Polynucleotide phosphorylase (PNPase) is an exoribonuclease and poly(A) polymerase postulated to function in the cytosol and mitochondrial matrix [8].
• PNPase is required for the optimal functioning of the Yersinia type three secretion system (TTSS), an organelle that injects effector proteins directly into host cells [3].
• Here, we show that PNPase also enhances the ability of Yersinia pseudotuberculosis and Yersinia pestis to withstand the killing activities of murine macrophages [3].
• The most potent inhibitor of human erythrocyte PNPase, [[[5-(2-amino-1,6-dihydro-6-oxo-9H-purin-9- yl)pentyl]phosphinico]methyl]phosphonic acid (2b), was a multisubstrate analogue inhibitor with a Ki' of 3.1 nM [12].

Associations of PNPT1 with chemical compounds

• The mutation responsible (pnp-71 ) has substituted a highly conserved glycine residue in the KH domain of PNPase with aspartate [10].
• These inhibitors displayed competitive kinetics with respect to inosine and inorganic phosphate, which showed that these compounds possess binding determinants for both the purine- and phosphate-binding domains of the enzyme, characteristics that are consistent with 3f and 3j being multisubstrate analogue inhibitors of PNPase [13].
• An initial set of six analogues of the weak PNPase inhibitor 9-benzylguanine (2) contained a phosphonic acid group linked to the ortho, meta, or para position of the aryl moiety via two- or three-atom spacers [13].
• The amine-phosphate interaction also served to confirm that a quinazolin-4(3H)-one binds in the PNPase active sites like a purine substrate [14].
• The reaction of 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole [NBD-Cl] with purified eel electrophax Na+ and K+ stimulated adenosine triphosphatase [(Na-K)ATPase] has been monitored by changes in the (Na-K)ATPase activity, the K+ stimulated p-nitrophenyl phosphatase [PNPase] activity, and the protein ultraviolet absorption spectrum [15].

Physical interactions of PNPT1

• We conclude from these studies that PNPase is the RNA-binding cofactor for this poly(A) polymerase and is an integral player in the reaction catalyzed by this enzyme [16].

Other interactions of PNPT1

• In contrast in HD PNPase, AMPA and CDA were increased (P less than 0-01) while ADA was in the normal range [17].
• In CLL a significant reduction (P less than 0-001) of AMPA, PNPase and ADA activities was observed without variation of CDA [17].
• Phosphonate acyclic derivates of guanines, pyrazolo[3,4-d]pyrimidines, and triazolo[4,5-d]-pyrimidines (8-azaguanines) are inhibitors of the enzyme purine nucleoside phosphorylase (PNPase) with Ki' values ranging from 0.05 to 1.6 microM [18].
• The enzyme activities were calculated by measurement of the decrease in the PNPase substrate, inosine, and the increase in the HGPRTase product, inosine-5'-monophosphate [19].
• Polynucleotide phosphorylase (PNPase) is a phosphate-dependent 3' to 5' exonuclease widely diffused among bacteria and eukaryotes [20].

Analytical, diagnostic and therapeutic context of PNPT1

• Electron microscopy shows the exosome to resemble PNPase but with key differences likely related to the position of RNA binding domains, and to the location of domains unique to the exosome [21].
• HPLC is highly suitable for PNPase as both the forward and reverse reactions can be monitored [22].
• Hypoxanthine-guanine phosphoribosyl transferase (HGPRTase) and purine nucleoside phosphorylase (PNPase) activities were simultaneously determined in erythrocyte lysates, using the reversed-phase mode of high-performance liquid chromatography [19].
• CI-972 (2,6-diamino-3,5-dihydro-7-(3-thienylmethyl)-4H-pyrrolo[3,2- d]pyrimidin-4-one monohydrochloride, monohydrate) is a competitive inhibitor of PNPase (E.C. 2.4.2.1., Ki = 0.83 microM) entering clinical trials as a T cell-selective immunosuppressive agent [23].
• In this work, we performed a systematic PCR mutagenesis of discrete pnp regions and screened the mutants for diverse phenotypic traits affected by PNPase [20].

References