E-Locus - Institutional Repository of the University of Crete

Home Search

Results - Details

Search command : Author="Περγαντής" And Author="Σπυρίδων"

Current Record: 16 of 56

[Add to Basket]

Identifier

000443253

Title

Μελέτη των πρωτεϊνικών συμπλόκων και των ισομορφών της πρωτεΐνης TP53 σε διαφορετικές κυτταρικές σειρές-μοντέλα του ανθρώπινου λεμφώματος πριν και μετά τη Ν3a επαγόμενη ενεργοποίησή της

Alternative Title

Study of protein complexes and isoforms of TP53 protein in different cell lines-models of human lymphoma before and after N3a induced activation

Author

Γεροπούλου, Νικολέτα Κ.

Thesis advisor

Αϊβαλιώτης, Μιχαήλ
Παυλίδης, Ιωάννης

Reviewer

Περγαντής, Σπυρίδων

Abstract

Lymphoma is the most common haematological malignancy affecting the cells of the immune system and is divided into two main groups, Hodgkin's (HL) and non-Hodgkin's lymphoma (NHL). In most cases of lymphoma, p53 is wild-type (wt) but is inactivated by an overexpressed protein, MDM2. The use of Nutlin-3a (chemical antagonist of MDM2) in lymphomas, mainly promotes apoptosis, through the reactivation of p53, a promising therapeutic strategy. However, there are at least 12 protein isoforms of p53 with differences in size and to a lesser extent in the amino acid sequence, with a still unclear role in lymphomas and p53 function. These isoforms have been found to determine and regulate the different actions of p53 (transcriptional regulation, protein interactions, etc.). Typical examples are p53β, which forms a protein complex with p53 and enhances its transcriptional activity, but also Δ133p53, in the presence of which the apoptotic effect of p53 is strongly inhibited. The aim of this study is to investigate the isoforms of wt-p53, as well as the detection of the protein complexes in which they are involved, in human lymphoma cells, against N3a induced activation of p53. For this purpose, model cell lines of human lymphoma were used. Through cell cultures the cells developed the desired number of cells and were incubated with and without N3a. Formaldehyde was then used in order to stabilize the protein interactions followed by cell lysis to isolate their proteins. After determination of their protein concentration with the BCA method, the protein extracts were immunoblotted by western blot, using antibodies to both p53 and its isoforms, as well as p21, which is the target of p53 activation and MDM2 which is its negative regulator. By special data processing, on the one hand the oligomerization of p53 before and after N3a, as well as its participation in complexes, was detected and quantified and on the other hand, the presence and relative abundance of different molecular weight isoforms of wt-p53 on lymphoma were evaluated in mantle cell lymphoma. The increase in oligomerization of p53 was confirmed after the addition of N3a, which is consistent with its tetramerification, but also the identification of p53 isoforms based on their molecular weight, but without being able to clearly distinguish isoforms α, β, γ. Isoforms α, β, γ will be further studied in future experiments using mass spectrometry.

Language

Greek

Subject

Immunoblotting

Isoforms

Lymphoma

MDM2

Mantle cell lymphoma

Nutlin-3a

p21

p53

Ανοσοαποτύπωση