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J. Biol. Chem., Vol. 275, Issue 47, 36803-36810, November 24, 2000
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From the Departments of We have studied the role of phosphatidylinositol
3-OH kinase (PI3K)-Akt signaling in transforming growth factor
Phosphatidylinositol 3-Kinase Function Is Required for
Transforming Growth Factor
-mediated Epithelial to Mesenchymal
Transition and Cell Migration*
§,§,§¶
**
Medicine,
¶ Cell Biology, and Pathology, Vanderbilt
University School of Medicine, ** Department of Veteran
Affairs Medical Center, and § Vanderbilt-Ingram Cancer
Center, Nashville Tennessee 37232
(TGF)-mediated epithelial to mesenchymal transition (EMT). In NMuMG
mammary epithelial cells, exogenous TGF1 induced phosphorylation of
Akt at Ser-473 and Akt in vitro kinase activity
against GSK-3 within 30 min. These responses were temporally
correlated with delocalization of E-cadherin, ZO-1, and integrin
1 from cell junctions and the acquisition of spindle
cell morphology. LY294002, an inhibitor of the p110 catalytic subunit
of PI3K, and a dominant-negative mutant of Akt blocked the
delocalization of ZO-1 induced by TGF1, whereas transfection of
constitutively active p110 induced loss of ZO-1 from tight junctions.
In addition, LY294002 blocked TGF-mediated C-terminal
phosphorylation of Smad2. Consistent with these data, TGF-induced
p3TP-Lux and p(CAGA)12-Lux reporter activities were inhibited by LY294002 and transiently expressed dominant-negative p85
and Akt mutants in NMuMG and 4T1 cells. Dominant-negative RhoA
inhibited TGF-induced phosphorylation of Akt at Ser-473, whereas
constitutively active RhoA increased the basal phosphorylation of Akt,
suggesting that RhoA in involved in TGF-induced EMT. Finally,
LY294002 and neutralizing TGF1 antibodies inhibited ligand-independent constitutively active Akt as well as basal and
TGF-stimulated migration in 4T1 and EMT6 breast tumor cells. Taken
together, these data suggest that PI3K-Akt signaling is required for
TGF-induced transcriptional responses, EMT, and cell migration.
*
This work was supported by Public Health Service
(PHS) Grant R01 CA62212, U. S. Department of Defense, U. S. Army Medical Research Material Command Grant DAMD17-98-1-8262, a
Clinical Investigator Award from the Department of Veterans Affairs (to
C. L. A.), PHS Grant R35 CA42572 (to H. L. M.), National
Institutes of Health Training Grant CA09592 (to N. A. B.), and
Vanderbilt-Ingram Cancer Center NCI National Institutes of Health
Support Grant CA68485.The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
To whom correspondence should be addressed: Div. of
Hematology-Oncology, Vanderbilt University School of Medicine, 22nd
Ave. South, 1956 TVC, Nashville, TN 37232-5536. E-mail:
carlos.arteaga@ mcmail.vanderbilt.edu.
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