Interrogation of the RP-MDM2-P53 Axis in Human Ribosomopathies
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CitationRaiser, David Michael. 2015. Interrogation of the RP-MDM2-P53 Axis in Human Ribosomopathies. Doctoral dissertation, Harvard University, Graduate School of Arts & Sciences.
AbstractRibosomopathies such as Diamond-Blackfan anemia (DBA) and 5q- syndrome are human diseases characterized by heterozygous loss or mutation of ribosome-associated genes. Hallmarks of these diseases include a macrocytic anemia in an otherwise normocellular bone marrow and, in some cases, developmental defects such as craniofacial or thumb abnormalities. Haploinsufficiency of ribosome-associated genes leads to dysfunctional ribosome biogenesis, which ultimately results in inhibition of MDM2 and consequent aberrant activation of the p53 pathway. Multiple genetic models have shown that p53 loss partially or completely rescues phenotypes associated with ribosomal protein (RP) haploinsufficiency, implicating p53 as the primary driver of those phenotypes in these diseases. However, the details of the molecular cascade leading to p53 activation – the RP-MDM2-p53 axis – are not fully understood. The thesis work presented here aimed to better characterize the molecular players downstream of ribosome dysfunction involved in modulation of ribosomopathy-associated phenotypes, as well as to identify novel therapeutic opportunities.
The proteomics screen characterized the protein binding partner profiles of MDM2 in cells with and without ribosome dysfunction. We found several RPs, including RPL5, RPL11, RPL23, and RPL38, to be commonly associated with MDM2, though RP deficiency may enhance the binding of RPL5 and RPL11. We also identified IGF1R as associated with MDM2 and selectively degraded in RP-deficient hematopoietic stem and progenitor cell (HSPC) cultures, and we showed that this protein loss contributes to defective erythropoiesis. In addition, a chemical screen identified calmodulin inhibitors as effective in rescuing ribosomopathy-associated phenotypes in zebrafish, mouse, and primary human HSPC models. We showed that these compounds (including the antipsychotic trifluoperazine) modulated p53 activity by inhibiting its nuclear localization, probably through inhibition of calmodulin-dependent CHK2. Lastly, we found that heterozygous RP gene deletion is a common feature of many human cancers. We further showed that silencing of a number of RPs frequently deleted in cancer results in p53 pathway activation, and that RP-deleted cancers have defects in ribosomal RNA processing. Together, this work adds novel insight to several aspects of ribosomopathy pathology and the RP-MDM2-p53 axis, and it further provides foundational evidence for novel therapeutic approaches to both ribosompathies and RP-deficient cancers.
Citable link to this pagehttp://nrs.harvard.edu/urn-3:HUL.InstRepos:17467484
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