DTYMK is essential for genome integrity and neuronal survival

Jo Vanoevelen ,Jörgen Bierau,Carlos R Ferreira,Liya Wang,Mark O’Driscoll,Marianna Bugiani,Junmei Hu Frisk,Erik‐Jan Kamsteeg ,Janine C Grashorn ,Richard J Rodenburg ,Debby M.e.i Hellebrekers ,Ghada M H Abdel-Salam ,Ellen Lambrichs,Levinus A Bok,Rita Colnaghi,Han G Brunner,Arthur Van Den Wijngaard,Marjo S Van Der Knaap,Ron A Wevers,Constance T R M Stumpel

doi:10.1007/s00401-021-02394-0

Abstract

Nucleotide metabolism is a complex pathway regulating crucial cellular processes such as nucleic acid synthesis, DNA repair and proliferation. This study shows that impairment of the biosynthesis of one of the building blocks of DNA, dTTP, causes a severe, early-onset neurodegenerative disease. Here, we describe two unrelated children with bi-allelic variants in DTYMK, encoding dTMPK, which catalyzes the penultimate step in dTTP biosynthesis. The affected children show severe microcephaly and growth retardation with minimal neurodevelopment. Brain imaging revealed severe cerebral atrophy and disappearance of the basal ganglia. In cells of affected individuals, dTMPK enzyme activity was minimal, along with impaired DNA replication. In addition, we generated dtymk mutant zebrafish that replicate this phenotype of microcephaly, neuronal cell death and early lethality. An increase of ribonucleotide incorporation in the genome as well as impaired responses to DNA damage were observed in dtymk mutant zebrafish, providing novel pathophysiological insights. It is highly remarkable that this deficiency is viable as an essential component for DNA cannot be generated, since the metabolic pathway for dTTP synthesis is completely blocked. In summary, by combining genetic and biochemical approaches in multiple models we identified loss-of-function of DTYMK as the cause of a severe postnatal neurodegenerative disease and highlight the essential nature of dTTP synthesis in the maintenance of genome stability and neuronal survival.

Highlights

Cellular processes such as proliferation and differentiation, DNA and RNA synthesis, repair and catabolism require a tightly regulated nucleotide metabolism
Brain pathology confirmed massive neuronal dropout in virtually the entire brain, sparing the brain stem. This striking neurodegenerative phenotype is observed in dtymk mutant zebrafish which exhibit microcephaly, brain edema and neuronal apoptosis (Fig. 3)
We present DTYMK variants in two unrelated families and provide biochemical data from fibroblasts of affected individuals and an additional animal model

Summary

Introduction

Cellular processes such as proliferation and differentiation, DNA and RNA synthesis, repair and catabolism require a tightly regulated nucleotide metabolism. Thymidine nucleotides are biosynthesized by reutilization of thymine and thymidine known as the salvage pathway (Fig. 1a). DTMP (deoxythymidine monophosphate) is formed by the de novo pathway, in which dUMP (deoxyuridine monophosphate) is converted to dTMP by thymidylate synthase (Fig. 1a). Exome analysis identified variants in DTYMK (deoxythymidine kinase) that segregate with this condition. DTYMK encodes dTMPK (deoxythymidine monophosphate kinase), a key enzyme in dTTP nucleotide metabolism (Fig. 1a). The thymidine de novo and salvage pathways converge at the point of the phosphorylation of dTMP to dTDP by dTMPK [13]. This makes dTMPK the bottleneck for dTTP biosynthesis (Fig. 1a)

Methods

Results

Conclusion