The role of the DNA damage response in zebrafish and cellular models of Diamond Blackfan anemia.

Nadia Danilova,Caius G Radu,David Nathanson,Elena Bibikova,Yoan Konto,Elizabeth Dimitrova,Todd M Covey,Bertil Glader,Anne Lindgren,Shuo Lin,Kathleen M Sakamoto

doi:10.1242/dmm.015495

Abstract

Ribosomal biogenesis involves the processing of pre-ribosomal RNA. A deficiency of some ribosomal proteins (RPs) impairs processing and causes Diamond Blackfan anemia (DBA), which is associated with anemia, congenital malformations and cancer. p53 mediates many features of DBA, but the mechanism of p53 activation remains unclear. Another hallmark of DBA is the upregulation of adenosine deaminase (ADA), indicating changes in nucleotide metabolism. In RP-deficient zebrafish, we found activation of both nucleotide catabolism and biosynthesis, which is consistent with the need to break and replace the faulty ribosomal RNA. We also found upregulation of deoxynucleotide triphosphate (dNTP) synthesis – a typical response to replication stress and DNA damage. Both RP-deficient zebrafish and human hematopoietic cells showed activation of the ATR/ATM-CHK1/CHK2/p53 pathway. Other features of RP deficiency included an imbalanced dNTP pool, ATP depletion and AMPK activation. Replication stress and DNA damage in cultured cells in non-DBA models can be decreased by exogenous nucleosides. Therefore, we treated RP-deficient zebrafish embryos with exogenous nucleosides and observed decreased activation of p53 and AMPK, reduced apoptosis, and rescue of hematopoiesis. Our data suggest that the DNA damage response contributes to p53 activation in cellular and zebrafish models of DBA. Furthermore, the rescue of RP-deficient zebrafish with exogenous nucleosides suggests that nucleoside supplements could be beneficial in the treatment of DBA.

Highlights

Ribosome biogenesis is the most energy-consuming process in the cell (Thomas, 2000)
The authors find that both the expression of enzymes involved in deoxynucleotide triphosphate (dNTP) biosynthesis and the incorporation of exogenous nucleotides into their DNA is increased in ribosomal proteins (RPs)-deficient zebrafish
These findings point to increased DNA repair. In accordance with this interpretation, the authors find that markers for DNA damage are upregulated in RP-deficient zebrafish and in human fetal liver cells that are deficient in RPS19

Summary

Introduction

Ribosome biogenesis is the most energy-consuming process in the cell (Thomas, 2000). It starts with the transcription of pre-ribosomal RNA (pre-rRNA) by RNA polymerase I (Kressler et al, 2010). Mutations in some RPs impair the processing of pre-rRNA, leading to the accumulation of defective rRNAs in cells (Choesmel et al, 2007; Flygare et al, 2007; Léger-Silvestre et al, 2005). It causes the Minute phenotype in Drosophila and Diamond-Blackfan anemia (DBA) syndrome in humans (Lipton and Ellis, 2009; Marygold et al, 2007). Several mechanisms of p53 activation that suggest a special checkpoint controlling ribosomal biogenesis have been proposed, but the exact mechanism remains unclear (Fumagalli and Thomas, 2011)

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Conclusion