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Title: On the ubiquitination and protein-protein interactions of p53
Author: Saliba, David George
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2007
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In this study p53F270A, p53R175H, and wild-type (wt) p53 were used as a panel to dissect how oligomeric protein conformation affects p53 ubiquitination and binding of other proteins to p53. p53F270A is hyperubiquitinated in cells as determined by p53-immunopreciptation and immunoblotting with an anti-ubiquitin antibody. Using a mass-spectrometric approach, eukaryotic elongation factor 1a (eefla) was identified as binding specifically to hyperubiquitinated p53F270A. In this study, using a one-strep tagged approach, the interactome of p53 was extended by defining stable cellular binding proteins to wild-type and mutant p53. The system faithfully recapitulates the enhanced ubiquitination pattern of the unfolded oncogenic forms of p53. Further, mutually exclusive binding partners were identified for wt and mutant p53: the wild-type protein interacted predominantly with MDM2; whilst the mutant interacted selectively with Heat shock protein 70 (Hsp70) and the TCP-1 beta chaperonin. Upon analyzing the effects of stresses on the p53-interactomes, we identified a Nutlin and 17-(allylamino)-17-demethoxy-geldanamycin (17-AAG)-induced binding to a novel spiced form of HSP90 lacking the N-terminus (HSP90ΔN). Ionizing radiation did not induce the formation of a p53: HSP90ΔN complex. By contrast, the oncogenic and hyperubiquitinated mutant p53 was constitutively bound to HSP90ΔN. These data expand on the Interactome of p53 and identify eefla and HSP90ΔN as novel components of signalling pathways that control p53 homeostasis.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available