title: A defective dNTP pool hinders DNA replication in cell cycle-reactivated terminally differentiated muscle cells.
abstract: Terminally differentiated cells are defined by their inability to proliferate. When forced to re-enter the cell cycle, they generally cannot undergo long-term replication. Our previous work with myotubes has shown that these cells fail to proliferate because of their intrinsic inability to complete DNA replication. Moreover, we have reported pronounced modifications of deoxynucleotide metabolism during myogenesis. Here we investigate the causes of incomplete DNA duplication in cell cycle-reactivated myotubes (rMt). We find that rMt possess extremely low levels of thymidine triphosphate (dTTP), resulting in very slow replication fork rates. Exogenous administration of thymidine or forced expression of thymidine kinase increases deoxynucleotide availability, allowing extended and faster DNA replication. Inadequate dTTP levels are caused by selective, differentiation-dependent, cell cycle-resistant suppression of genes encoding critical synthetic enzymes, chief among which is thymidine kinase 1. We conclude that lack of dTTP is at least partially responsible for the inability of myotubes to proliferate and speculate that it constitutes an emergency barrier against unwarranted DNA replication in terminally differentiated cells.
authors.affiliation: Department of Cell Biology and Neurosciences, Italian National Institute of Health, Rome, Italy.
authors.forename: Deborah
authors.initials: D
authors.lastname: Pajalunga
authors.affiliation: Department of Biology, University of Padua, Padua, Italy.
authors.forename: Elisa
authors.initials: E
authors.lastname: Franzolin
authors.affiliation: Department of Biology, University of Padua, Padua, Italy.
authors.forename: Martina
authors.initials: M
authors.lastname: Stevanoni
authors.affiliation: Department of Cell Biology and Neurosciences, Italian National Institute of Health, Rome, Italy.
authors.forename: Sara
authors.initials: S
authors.lastname: Zribi
authors.affiliation: Department of Cell Biology and Neurosciences, Italian National Institute of Health, Rome, Italy.
authors.forename: Nunzia
authors.initials: N
authors.lastname: Passaro
authors.affiliation: Department of Research, Advanced Diagnostics and Technological Innovation, SAFU Unit, Translational Research Area, Regina Elena National Cancer Institute, Rome, Italy.
authors.forename: Aymone
authors.identifier: 0000-0002-7661-9059
authors.initials: A
authors.lastname: Gurtner
authors.affiliation: Department of Cell Biology and Neurosciences, Italian National Institute of Health, Rome, Italy.
authors.forename: Samantha
authors.initials: S
authors.lastname: Donsante
authors.affiliation: Department of Cell Biology and Neurosciences, Italian National Institute of Health, Rome, Italy.
authors.forename: Daniela
authors.initials: D
authors.lastname: Loffredo
authors.affiliation: Department of Cell Biology and Neurosciences, Italian National Institute of Health, Rome, Italy.
authors.forename: Lidia
authors.initials: L
authors.lastname: Losanno
authors.affiliation: Department of Biology, University of Padua, Padua, Italy.
authors.forename: Vera
authors.initials: V
authors.lastname: Bianchi
authors.affiliation: Department of Biology, University of Padua, Padua, Italy.
authors.forename: Antonella
authors.identifier: 0000-0001-6691-257X
authors.initials: A
authors.lastname: Russo
authors.affiliation: Department of Biology, University of Padua, Padua, Italy.
authors.forename: Chiara
authors.initials: C
authors.lastname: Rampazzo
authors.affiliation: Department of Cell Biology and Neurosciences, Italian National Institute of Health, Rome, Italy.
authors.forename: Marco
authors.initials: M
authors.lastname: Crescenzi
mesh_terms: D000818:Animals; D002453:Cell Cycle / Q000187:drug effects* / Q000235:genetics; D002454:Cell Differentiation / Q000235:genetics; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics / Q000378:metabolism; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000235:genetics / Q000378:metabolism; D004261:DNA Replication / Q000187:drug effects*; D003845:Deoxycytosine Nucleotides / Q000378:metabolism; D005786:Gene Expression Regulation; D006657:Histones / Q000235:genetics / Q000378:metabolism; D051379:Mice; D024510:Muscle Development / Q000235:genetics; D018485:Muscle Fibers, Skeletal / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D061251:Primary Cell Culture; D032496:Satellite Cells, Skeletal Muscle / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D013936:Thymidine / Q000494:pharmacology*; D013937:Thymidine Kinase / Q000235:genetics* / Q000378:metabolism; D013938:Thymidine Monophosphate / Q000378:metabolism; D013942:Thymine Nucleotides / Q000172:deficiency*
chemical_list: D050759:Cyclin-Dependent Kinase Inhibitor p21; D003845:Deoxycytosine Nucleotides; D006657:Histones; D013942:Thymine Nucleotides; C475961:gamma-H2AX protein, mouse; D050760:Cyclin-Dependent Kinase Inhibitor p27; C024107:2'-deoxycytidine 5'-triphosphate; D013938:Thymidine Monophosphate; C105339:thymidine kinase 2; D013937:Thymidine Kinase; C092717:thymidine kinase 1; C024157:thymidine 5'-triphosphate; D013936:Thymidine