INVESTIGADORES
ROSEMBLIT Cinthia
congresos y reuniones científicas
Título:
Signal transducer and activator of transcription 3 (Stat3) enhances Progesterone Receptor (PR) activation of transcription in breast cancer cells
Autor/es:
PORIETTI CJ ; ROSEMBLIT C ; BEGUELIN W ; DIAZ FLAQUE MC; RIVAS MA ; TKACH M ; CHARREAU EH ; SCHILLACI R ; ELIZALDE PV
Lugar:
Carefree, Arizona
Reunión:
Congreso; FASEB Summer Conferences. Extranuclear Steroid Receptors: Integration with Multiple Signaling Pathways; 2008
Institución organizadora:
FASEB
Resumen:
Signal transducer and activator of transcription 3 (Stat3) enhances
Progesterone Receptor (PR) activation of transcription in breast cancer
cells.
We have previously demonstrated that the synthetic progestin
medroxyprogesterone acetate (MPA) is able to induce Stat3
transcriptional
activation, which is in turn an absolute requirement for progestin
stimulation of
in vitro and in vivo breast cancer growth. We have performed our study
both in
C4HD cells from an experimental model of hormonal carcinogenesis in
which
MPA induced mammary adenocarcinomas in Balb/c mice, and in the human
breast cancer cell line T47D (Mol Cell Biol, 25: 4826, 2005).Therefore,
we
here assessed whether the requirement of Stat3 activation in progestin-
stimulated breast cancer cell proliferation could be due to
bi-directional cross-
talks between progestins and Stat3, where Stat3 in turn regulates PR
transcriptional activation. We studied whether Stat3, acting as a
coactivator,
could modulate PR function, both when PR binds to specific progesterone
response elements (PRE) in the promoter regions of target genes, such as
bcl-X gene, and when PR regulates the transcription of p21 gene, which
lacks canonical PREs in its promoter region. Our present findings
evidenced
that MPA treatment of C4HD cells induced an increase in bcl-XL mRNA
levels. This effect was completely abolished by transfection with a
dominant
negative Stat3 expression vector, Stat3Y705-F, and in the presence of
the
progestin antagonist RU486. In addition, we also found that MPA
treatment of
both C4HD and T47D cells, induced an increase in the levels of p21
protein
expression, which was further enhanced in a dose-dependent manner in the
presence of a plasmid expressing a constitutively activated Stat3
mutant,
Stat3-C. To study the effect of Stat3 on PR-mediated transcriptional
activity,
C4HD and T47D cells were transiently transfected with a luciferase
reporter
plasmid under the control of the bcl-XP4 promoter, together with
Stat3-C.
MPA treatment resulted in an increase in luciferase activity, in
accordance
with previous results describing the presence of two hormone responsive
elements present in the fourth promoter of bcl-X gene. We found that
Stat3-C
enhanced progestin-induced PR transcriptional activity in a
dose-dependent
manner and that this effect was abolished by RU486 and by cotransfection
with Stat3Y705-F. Similar results were obtained with the
well-characterized
progesterone-responsive luciferase reporter plasmid MMTV-luc. We also
studied the effect of Stat3 on PR capacity to regulate p21 transcription
by
tethering to DNA-bound trans-acting factor Sp1. C4HD and T47D cells were
transiently transfected with a p21 promoter reporter construct
containing two
Sp1 binding sites, together with increasing amounts of Stat3-C. We found
that Stat3-C enhanced progestin-induced PR transcriptional activity in a
dose-
dependent manner. This effect was completely abolished when Stat3
expression was silenced using an siRNA strategy. Finally, we explored
the
specific association of Stat3 and PR to the PRE region of the bcl-X gene
in
the context of living cells, by performing Chromatin Immunoprecipitation
(ChIP) and Sequential ChIP Assays. Our findings unraveled that MPA
treatment of primary cultures of C4HD cells induced PR and Stat3
recruitment to the bcl-X promoter. We also evaluated the specific
association
of Stat3 and PR to the Sp-1 binding site in the p21 promoter and found
that
MPA treatment of T47D cells induced Sp-1, PR and Stat3 recruitment to
the
p21 promoter.
These results provide the first evidence that Stat3 acts as a
coactivator of PR
when PR binds directly to target genes containing PREs in the promoter
region, or when PR regulates transcription by tethering to Sp1.