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TGF-β induced TMEPAI/PMEPA1 inhibits canonical Smad signaling through R-Smad sequestration and promotes non-canonical PI3K/Akt signaling by reducing PTEN in triple negative breast cancer.

Singha PK, Pandeswara S, Geng H, Lan R, Venkatachalam MA, Saikumar P - Genes Cancer (2014)

Bottom Line: Firstly, TMEPAI binds and sequesters regulatory Smads2/3 and thereby decreases growth suppressive signaling by TGF-β.Moreover, signaling alterations produced by high TMEPAI were associated with oncogenic Snail expression and lung metastases.Together, our findings suggest that TMEPAI has dually critical roles to promote TGF-β dependent cancer cell growth and metastasis.

View Article: PubMed Central - PubMed

Affiliation: From the Department of Pathology, UT Health Science Center at San Antonio, TX.

ABSTRACT
TMEPAI (transmembrane prostate androgen-induced) is amplified at genomic, transcript and protein levels in triple-negative breast cancers and promotes TGF-β dependent growth, motility and invasion. Tumor promotion by TMEPAI depends on two different but related actions on TGF-β signaling. Firstly, TMEPAI binds and sequesters regulatory Smads2/3 and thereby decreases growth suppressive signaling by TGF-β. Secondly, increased expression of TMEPAI decreases PTEN (phosphatase and tensin homolog) abundance, and thereby increases TGF-β dependent tumor promotive PI3K/Akt signaling. These actions of TMEPAI give rise to increased cell proliferation and motility. Moreover, signaling alterations produced by high TMEPAI were associated with oncogenic Snail expression and lung metastases. Finally, an inverse correlation between TMEPAI and PTEN levels was confirmed in triple negative breast cancer tumor samples. Together, our findings suggest that TMEPAI has dually critical roles to promote TGF-β dependent cancer cell growth and metastasis. Thus, redirected TGF-β signaling through TMEPAI may play a pivotal role in TGF-β mediated tumor promotion.

No MeSH data available.


Related in: MedlinePlus

Smad requirement in TMEPAI expressionA) TMEPAI expression in three different triple negative breast cancer cell lines without or with TGF-β for 24h. B) Relative expressions of TMEPAI and Smad4 in MDA-MB-231 and MDA-MB-468 cells. C) Exogenous expression of Smad4 using a retroviral vector in MDA-MB-468 cells restores TMEPAI expression and TGF-β mediated induction of TMEPAI in MDA-MB-468 cells.
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Figure 2: Smad requirement in TMEPAI expressionA) TMEPAI expression in three different triple negative breast cancer cell lines without or with TGF-β for 24h. B) Relative expressions of TMEPAI and Smad4 in MDA-MB-231 and MDA-MB-468 cells. C) Exogenous expression of Smad4 using a retroviral vector in MDA-MB-468 cells restores TMEPAI expression and TGF-β mediated induction of TMEPAI in MDA-MB-468 cells.

Mentions: TMEPAI protein in MDA-MB-231 triple negative breast cancer cells is induced by TGF-β in a concentration dependent manner as determined by exposing cells to graded amounts of TGF-β (0.1 0.25, 0.5, 1, 1.5 and 2 ng per ml) for 24 h. The induction was seen even at 100 pg/ml of TGF-β (2.5 fold) and reached maximum (8.4 fold) at 1 ng/ ml of TGFβ concentration (Fig.1B). The TMEPAI protein induction appeared to be rapid that occurred within 30 min and reached peak levels after 3 h of exposure to 1 ng/ml TGF-β (Fig.1C). Induction of TMEPAI was blocked by both transcriptional and translational inhibitors such as actinomycin D and cycloheximide, respectively (Supplemental Fig.2A). While MDA-MB-231 cancer cells are resistant to TGF-β mediated growth inhibition (Fig.1D), normal HMEC cells are sensitive (Fig.1E) to such inhibition. In some cancer cells like MDA-MB-231, TGF-β may even further stimulate their growth (see Fig.1D). Inclusion of TGF-β signaling inhibitor SB431542 in the growth medium blocked the proliferation of MDA-MB-231 cancer cells without any effect on the HMEC growth suggesting that MDA-MB-231 cells are dependent on autocrine TGF-β signaling for their proliferation but not HMEC (see Figs. 1D & 1E). Correspondingly, basal levels of TMEPAI were detected in MDA-MB-231 cancer cells, but not in normal mammary epithelial cells, HMEC (Figs.1D &1E). Importantly, TMEPAI induction by TGF-β was rather marginal in HMEC, while abundant in MDA-MB-231 cells [see Figs. 1D & 1E, insets]. TGF-β receptor I kinase inhibitor SB431542, inhibited both basal and TGF-β induced expression of TMEPAI protein (Figs.1D &1E, insets) and mRNA in MDA-MB-231 cells (Supplemental Fig. 2B), indicating positive regulation of TMEPAI expression by TGF-β. Moreover, TGF-β is much stronger than other growth factors such as EGF (epidermal growth factor) or growth factor like lipid LPA (lysophasphatidic acid) in inducing TMEPAI (Supplemental Fig.2C) suggesting the specificity for TGF-β. In addition, TMEPAI was similarly induced by TGF-β in other TNBC cell lines BT-20 and HCC1937, except in MDA-MB-468 cells (Fig.2A), because these cells lacked Smad4 expression required for TGF-β signaling (Fig.2B). In fact, exogenous expression of Smad4 in MDA-MB-468 cells restored induction of TMEPAI by TGF-β (Fig.2C) further confirming that TGF-β signaling is required for TMEPAI induction.


TGF-β induced TMEPAI/PMEPA1 inhibits canonical Smad signaling through R-Smad sequestration and promotes non-canonical PI3K/Akt signaling by reducing PTEN in triple negative breast cancer.

Singha PK, Pandeswara S, Geng H, Lan R, Venkatachalam MA, Saikumar P - Genes Cancer (2014)

Smad requirement in TMEPAI expressionA) TMEPAI expression in three different triple negative breast cancer cell lines without or with TGF-β for 24h. B) Relative expressions of TMEPAI and Smad4 in MDA-MB-231 and MDA-MB-468 cells. C) Exogenous expression of Smad4 using a retroviral vector in MDA-MB-468 cells restores TMEPAI expression and TGF-β mediated induction of TMEPAI in MDA-MB-468 cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 2: Smad requirement in TMEPAI expressionA) TMEPAI expression in three different triple negative breast cancer cell lines without or with TGF-β for 24h. B) Relative expressions of TMEPAI and Smad4 in MDA-MB-231 and MDA-MB-468 cells. C) Exogenous expression of Smad4 using a retroviral vector in MDA-MB-468 cells restores TMEPAI expression and TGF-β mediated induction of TMEPAI in MDA-MB-468 cells.
Mentions: TMEPAI protein in MDA-MB-231 triple negative breast cancer cells is induced by TGF-β in a concentration dependent manner as determined by exposing cells to graded amounts of TGF-β (0.1 0.25, 0.5, 1, 1.5 and 2 ng per ml) for 24 h. The induction was seen even at 100 pg/ml of TGF-β (2.5 fold) and reached maximum (8.4 fold) at 1 ng/ ml of TGFβ concentration (Fig.1B). The TMEPAI protein induction appeared to be rapid that occurred within 30 min and reached peak levels after 3 h of exposure to 1 ng/ml TGF-β (Fig.1C). Induction of TMEPAI was blocked by both transcriptional and translational inhibitors such as actinomycin D and cycloheximide, respectively (Supplemental Fig.2A). While MDA-MB-231 cancer cells are resistant to TGF-β mediated growth inhibition (Fig.1D), normal HMEC cells are sensitive (Fig.1E) to such inhibition. In some cancer cells like MDA-MB-231, TGF-β may even further stimulate their growth (see Fig.1D). Inclusion of TGF-β signaling inhibitor SB431542 in the growth medium blocked the proliferation of MDA-MB-231 cancer cells without any effect on the HMEC growth suggesting that MDA-MB-231 cells are dependent on autocrine TGF-β signaling for their proliferation but not HMEC (see Figs. 1D & 1E). Correspondingly, basal levels of TMEPAI were detected in MDA-MB-231 cancer cells, but not in normal mammary epithelial cells, HMEC (Figs.1D &1E). Importantly, TMEPAI induction by TGF-β was rather marginal in HMEC, while abundant in MDA-MB-231 cells [see Figs. 1D & 1E, insets]. TGF-β receptor I kinase inhibitor SB431542, inhibited both basal and TGF-β induced expression of TMEPAI protein (Figs.1D &1E, insets) and mRNA in MDA-MB-231 cells (Supplemental Fig. 2B), indicating positive regulation of TMEPAI expression by TGF-β. Moreover, TGF-β is much stronger than other growth factors such as EGF (epidermal growth factor) or growth factor like lipid LPA (lysophasphatidic acid) in inducing TMEPAI (Supplemental Fig.2C) suggesting the specificity for TGF-β. In addition, TMEPAI was similarly induced by TGF-β in other TNBC cell lines BT-20 and HCC1937, except in MDA-MB-468 cells (Fig.2A), because these cells lacked Smad4 expression required for TGF-β signaling (Fig.2B). In fact, exogenous expression of Smad4 in MDA-MB-468 cells restored induction of TMEPAI by TGF-β (Fig.2C) further confirming that TGF-β signaling is required for TMEPAI induction.

Bottom Line: Firstly, TMEPAI binds and sequesters regulatory Smads2/3 and thereby decreases growth suppressive signaling by TGF-β.Moreover, signaling alterations produced by high TMEPAI were associated with oncogenic Snail expression and lung metastases.Together, our findings suggest that TMEPAI has dually critical roles to promote TGF-β dependent cancer cell growth and metastasis.

View Article: PubMed Central - PubMed

Affiliation: From the Department of Pathology, UT Health Science Center at San Antonio, TX.

ABSTRACT
TMEPAI (transmembrane prostate androgen-induced) is amplified at genomic, transcript and protein levels in triple-negative breast cancers and promotes TGF-β dependent growth, motility and invasion. Tumor promotion by TMEPAI depends on two different but related actions on TGF-β signaling. Firstly, TMEPAI binds and sequesters regulatory Smads2/3 and thereby decreases growth suppressive signaling by TGF-β. Secondly, increased expression of TMEPAI decreases PTEN (phosphatase and tensin homolog) abundance, and thereby increases TGF-β dependent tumor promotive PI3K/Akt signaling. These actions of TMEPAI give rise to increased cell proliferation and motility. Moreover, signaling alterations produced by high TMEPAI were associated with oncogenic Snail expression and lung metastases. Finally, an inverse correlation between TMEPAI and PTEN levels was confirmed in triple negative breast cancer tumor samples. Together, our findings suggest that TMEPAI has dually critical roles to promote TGF-β dependent cancer cell growth and metastasis. Thus, redirected TGF-β signaling through TMEPAI may play a pivotal role in TGF-β mediated tumor promotion.

No MeSH data available.


Related in: MedlinePlus