Mitochondrial DNA depletion and thymidine phosphate pool dynamics in a cellular model of mitochondrial neurogastrointestinal encephalomyopathy

Giovanna Pontarin, Paola Ferraro, Maria L. Valentino, Michio Hirano, Peter Reichard, Vera Bianchi

Research output: Contribution to journalArticle

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Abstract

Mitochondrial (mt) neurogastrointestinal encephalomyopathy (MNGIE) is an autosomal recessive disease associated with depletion, deletions, and point mutations of mtDNA. Patients lack a functional thymidine phosphorylase and their plasma contains high concentrations of thymidine and deoxyuridine; elevation of the corresponding triphosphates probably impairs normal mtDNA replication and repair. To study metabolic events leading to MNGIE we used as model systems skin and lung fibroblasts cultured in the presence of thymidine and/or deoxyuridine at concentrations close to those in the plasma of the patients, a more than 100-fold excess relative to controls. The two deoxynucleosides increased the mt and cytosolic dTTP pools of skin fibroblasts almost 2-fold in cycling cells and 8-fold in quiescent cells. During up to a two-month incubation of quiescent fibroblasts with thymidine (but not with deoxyuridine), mtDNA decreased to ∼50% without showing deletions or point mutations. When we removed thymidine, but maintained the quiescent state, mtDNA recovered rapidly. With thymidine in the medium, the dTTP pool of quiescent cells turned over rapidly at a rate depending on the concentration of thymidine, due to increased degradation and resynthesis of dTMP in a substrate (=futile) cycle between thymidine kinase and 5′-deoxyribonucleotidase. The cycle limited the expansion of the dTTP pool at the expense of ATP hydrolysis. We propose that the substrate cycle represents a regulatory mechanism to protect cells from harmful increases of dTTP. Thus MNGIE patients may increase their consumption of ATP to counteract an unlimited expansion of the dTTP pool caused by circulating thymidine.

Original languageEnglish
Pages (from-to)22720-22728
Number of pages9
JournalJournal of Biological Chemistry
Volume281
Issue number32
DOIs
Publication statusPublished - Aug 11 2006

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Thymine Nucleotides
Mitochondrial Encephalomyopathies
Mitochondrial DNA
Thymidine
Deoxyuridine
Fibroblasts
Sequence Deletion
Point Mutation
Skin
Adenosine Triphosphate
Substrate Cycling
Thymidine Phosphorylase
Plasmas
Thymidine Kinase
Substrates
Hydrolysis
Repair
thymidine 5'-triphosphate
Degradation
Lung

ASJC Scopus subject areas

  • Biochemistry

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Mitochondrial DNA depletion and thymidine phosphate pool dynamics in a cellular model of mitochondrial neurogastrointestinal encephalomyopathy. / Pontarin, Giovanna; Ferraro, Paola; Valentino, Maria L.; Hirano, Michio; Reichard, Peter; Bianchi, Vera.

In: Journal of Biological Chemistry, Vol. 281, No. 32, 11.08.2006, p. 22720-22728.

Research output: Contribution to journalArticle

Pontarin, Giovanna ; Ferraro, Paola ; Valentino, Maria L. ; Hirano, Michio ; Reichard, Peter ; Bianchi, Vera. / Mitochondrial DNA depletion and thymidine phosphate pool dynamics in a cellular model of mitochondrial neurogastrointestinal encephalomyopathy. In: Journal of Biological Chemistry. 2006 ; Vol. 281, No. 32. pp. 22720-22728.
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