p53-Independent Cell Cycle and Erythroid Differentiation Defects in Murine Embryonic Stem Cells Haploinsufficient for Diamond Blackfan Anemia-Proteins: RPS19 versus RPL5

Abstract

Diamond Blackfan anemia (DBA) is a rare inherited bone marrow failure syndrome caused by ribosomal protein haploinsufficiency. DBA exhibits marked phenotypic variability, commonly presenting with erythroid hypoplasia, less consistently with non-erythroid features. The p53 pathway, activated by abortive ribosome assembly, is hypothesized to contribute to the erythroid failure of DBA. We studied murine embryonic stem (ES) cell lines harboring a gene trap mutation in a ribosomal protein gene, either Rps19 or Rpl5. Both mutants exhibited ribosomal protein haploinsufficiency and polysome defects. Rps19 mutant ES cells showed significant increase in p53 protein expression; however, there was no similar increase in the Rpl5 mutant cells. Embryoid body formation was diminished in both mutants but nonspecifically rescued by knockdown of p53. When embryoid bodies were further differentiated to primitive erythroid colonies, both mutants exhibited a marked reduction in colony formation, which was again nonspecifically rescued by p53 inhibition. Cell cycle analyses were normal in Rps19 mutant ES cells, but there was a significant delay in the G2/M phase in the Rpl5 mutant cells, which was unaffected by p53 knockdown. Concordantly, Rpl5 mutant ES cells had a more pronounced growth defect in liquid culture compared to the Rps19 mutant cells. We conclude that the defects in our RPS19 and RPL5 haploinsufficient mouse ES cells are not adequately explained by p53 stabilization, as p53 knockdown appears to increase the growth and differentiation potential of both parental and mutant cells. Our studies demonstrate that gene trap mouse ES cells are useful tools to study the pathogenesis of DBA.

Marrow failure: a window into ribosome biology

“…Intriguingly, depletion of RPL5 or RPL11 impairs cell-cycle progression in a p53-independent manner. 33,34 Mouse 21 and zebrafish 35 animal models also provide evidence for both p53-dependent and p53-independent pathways in DBA. Indeed, p53 activation appears to be a common downstream stress response pathway activated in response to a variety of different mechanisms underlying nonribosomal marrow failure syndromes such as anemia.…”

Clinical features, mutations and treatment of 104 patients of Diamond-Blackfan anemia in China: a single-center retrospective study

“…Both P53 dependent and independent pathways were demonstrated to participate in the pathogenesis of DBA [10,11]. To date, up to 70 % of the patients have a known mutation or deletion in their ribosomal proteins.…”

L‐Leucine improves the anaemia in models of Diamond Blackfan anaemia and the 5q‐ syndrome in a TP53‐independent way

“…However, rescue of RP-deficient anaemia in a TP53-null setting is rarely complete (Singh, et al 2014, Yadav, et al 2014), leading to the hypothesis that RP-deficient anaemia has a TP53-independent component. Furthermore, TP53 activation fails to account for several aspects of disease pathophysiology.…”