Lineage-Specific Activation of p53 in Response to Ribosomal Haploinsufficiency in Human Bone Marrow Cells.

Abstract 948

Haploinsufficiency for the ribosomal protein genes RPS14 and RPS19 have been implicated in the erythroid defect in the 5q- syndrome and Diamond Blackfan Anemia, respectively. However, the mechanism by which defective ribosome biogenesis causes erythroid failure is unknown. In this study, we found that shRNA mediated knockdown of RPS14 or RPS19 in primary human CD34+ cells stabilize TP53 by day 4 after infection with concomitant arrest of these cells at G1 stage of cell cycle. The levels of TP53 attained are comparable to the levels observed following gamma irradiation (5Gy) of the CD34+ cells. Using quantitative PCR, we confirmed that stabilized TP53 activates expression of downstream target genes MDM2, p21, Bax and Wig-1. Furthermore, treatment of the CD34+ cells with Nutlin-3 phenocopies RPS14 or RPS19 knockdown, suggesting that the mechanism of TP53 activation is mediated by MDM2 pathway. Conversely, treatment with pifithrin-alpha, which inhibits the transactivation activity of TP53, rescues the effects of RPS14 or RPS19 knockdown. The in vitro activation of TP53 in CD34+ cells was restricted to erythroid cell lineage, consistent with the clinical phenotype of RPS14 or RPS19 haploinsufficiency. Moreover, immunohistochemical analysis of bone marrow biopsies from patient with the 5q- syndrome demonstrated intense staining of TP53 that was restricted to erythroid progenitor cells. Taken together our study indicates that inhibition of ribosomal biogenesis causes TP53 activation selectively in erythroid progenitor cells. Clinically, TP53 staining of patient samples could be used as a diagnostic marker for some types of MDS.