Visit our Expo - Redox and Inflammation signaling 2012

Session XVII : Cell signaling in health and disease Poster XVII, 30
15-Deoxy-&12,14-prostaglandin J2 covalently modifies and functionally inactivates p53
in human breast cancer (MCF-7) cells
Do-Hee Kim, Hye-Kyung Na and Young-Joon Surh
National Research Laboratory of Molecular Carcinogenesis and Chemoprevention,
College of Pharmacy, Seoul National University, Seoul 151-742, Republic of Korea
Cyclopentenone prostaglandins (cyPGs) exert diverse cellular functions, such as antiinflammatory
and cytoprotective effects. 15-Deoxy- &12,14-prostaglandin J2 (15d-PGJ2), a
representative cyPG of the J series, has been reported to exert biphasic effects on proliferation
and apoptosis. In the present study, we found that MCF-7 human breast cancer cells treated
with 15d-PGJ2 accumulated the tumor suppressor gene product p53 in both cytosolic and
nuclear fractions. Under the same experimental conditions, however, the mRNA level of p53
remained unchanged. This prompted us to investigate the molecular mechanisms responsible
for the 15d-PGJ2-mediated accumulation of p53 protein. Murine double-minute 2 (MDM2), a
zinc finger protein, is known to negatively regulate p53. 15d-PGJ2 treatment caused failure of
proper p53-MDM2 interaction in MCF-7 cells which could stabilize p53 protein by blocking
MDM2-mediated degradation. We observed that 15d-PGJ2 covalently modify cysteine
residues of p53 in MCF-7 cells. Covalent modification of p53 by 15d-PGJ2 is considered to
interfere with DNA binding of p53. In support of this assumption, 15d-PGJ2 inhibited the
DNA binding ability of both recombinant p53 and MCF-7 nuclear extracts in a concentrationrelated
manner. Likewise, p53 conjugation with 15d-PGJ2 may hamper its interaction with
MDM2, and subsequent degradation. The majority of accumulated p53 in 15d-PGJ2-treated
MCF-7 cells represents a functionally inactive or mutant form as assessed by
immunoprecipitation and immunofluorescence analysis. It is anticipated that the electrophilic
carbon center located in the alpha,#-unsaturated carbonyl moiety of the cyclopentenone ring
might be critical for the 15d-PGJ2-induced covalent modification of p53, cysteine thiol,
which retarded its degradation. MCF-7 cells treated with 9,10-dihydro-15-deoxy-&12,14prostaglandin
J2 that lacks the electrophilic alpha #-unsaturated functionality failed to
accumulate p53 protein, lending further support to the above assumption. In summary, 15d-
PGJ2 directly conjugates with cysteine thiol residues of p53, interfering its interaction with
MDM2, thereby stabilizing p53. Covalent modification of p53 by 15d-PGJ2 also hampers p53
DNA binding, leading to functional inactivation of this tumor suppressor.
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