Identification of the interacting domain of p53 family members with P33ING1 protein
Author(s)
Advisor
Çetin-Atalay, RengülDate
2002Publisher
Bilkent University
Language
English
Type
ThesisItem Usage Stats
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Abstract
p53 is a tumor suppressor gene, which is mutated, in about 50% of human
cancers. The product of p53 gene encodes a sequence specific transcription factor.
The genes transactivated by p53 code for proteins that are implicated in the negative
regulation of cell proliferation and DNA damage repair. Two proteins, p63 and p73
members of p53 family, show striking homology to p53. p53 protein interacts with
several viral and cellular proteins and these interactions are important in the
regulation and dysregulation of the functions of p53. Another gene named, ING1 was
identified as a candidate tumor suppressor gene due to its functions in apoptosis and
cell cycle arrest. p24ING1, one of the protein product of ING1, was shown to enhance
the growth suppressor functions of p53. Furthermore a physical association between
p53 and p33ING1, another ING1 transcript, proteins has been detected by
immunoprecipitation. In this study, we investigated the physical interaction between
p53 family proteins and p33ING1 using in vitro techniques in order to determine the
region(s) of p53 family proteins and p33ING1 that enabled this interaction. As a
preliminary step for the study, the wild-type p53 cDNA and its several deletion
mutant constructs were used in GST pulldown assays and Far Western assays with
purified GST-p33 protein to map the interacting region on p53 protein. New deletion
mutant constructs of p53 protein were created and cloned into expression vectors for
the detailed analysis of the interacting domain of p53 protein. Also the other members
of p53 protein family, p63 and p73 were examined in vitro for interaction with
p33ING1. Deletion mutants of these proteins were created and cloned into expression
vectors for protein-protein interaction assays. The results of this study shows that p53
protein interacts with p33ING1 and suggests that oligomerization domain of p53
protein is needed for this interaction. In addition, for the first time, it was showed that
p63 and p73 proteins interact with p33ING1 and in p63 the C-terminus region is the
primary determinant region, involved in these interactions with p33ING1.