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Nucleoside Salvage Kinases Regulate Hematopoiesis by Linking Nucleotide Metabolism with DNA Replication Stress

  • Author(s): Austin, Wayne Robert
  • Advisor(s): Radu, Caius G
  • Czernin, Johannes
  • et al.
Abstract

Deoxycytidine kinase (dCK) is a rate-limiting enzyme in the salvage pathway of deoxribonucleotide triphosphate (dNTP) biosynthesis. dCK is highly expressed in hematopoietic tissues such as the bone marrow and thymus, indicating that the enzyme is important for the generation of a normal hematopoietic system. However, it has never been determined as to why dCK is highly expressed in these proliferating cell types versus others in the body. To begin answering this fundamental biological question, we generated mice that were globally deficient for dCK activity (dCK-/-) using genetic knockout technology. We found that inactivation of dCK activity results in severely inhibited differentiation of major hematopoietic lineages consisting of T cells, B cells, and erythroid cells. Examination of the cell cycle kinetics in dCK-/- cells from these distinct lineages revealed that these cells undergo cell cycle arrest early during the process of DNA replication in vivo. This arrest is associated with the presence of active DNA damage response, thus indicating that hematopoietic progenitor cells are subject to significant DNA replication stress in vivo. We found that induction of DNA replication stress in dCK-/- hematopoietic progenitors was dependent upon the salvage of thymidine by the nucleoside salvage enzyme thymidine kinase 1 (TK1). We subsequently demonstrated that simultaneous inactivation of both dCK and TK1 in mice greatly ameliorated the proliferation and differentiation defects that were present in hematopoietic progenitor cells from dCK-/- mice. The results from our studies thus demonstrate that dCK functions to counteract the DNA replication stress-inducing activity of TK1 that occurs during in vivo proliferation of murine hematopoietic progenitors.

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