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PKN1 modulates TGFβ and EGF signaling in HEC-1-A endometrial cancer cell line.

Attarha S, Saini RK, Andersson S, Mints M, Souchelnytskyi S - Onco Targets Ther (2014)

Bottom Line: The response of cells to TGFβ and EGF is mediated by a network of various intracellular regulators.It was observed that phosphorylation of Smad2, FAK, and Erk1/2 correlated with responses of the cells to TGFβ1 and EGF.PKN1 modulates TGFβ- and EGF-dependent regulation of cell proliferation, migration, and invasiveness, and therefore is a component of the network signaling downstream of TGFβ and EGF.

View Article: PubMed Central - PubMed

Affiliation: Department of Oncology-Pathology, Karolinska Institutet, Stockholm, Sweden ; Department of Women's and Children's Health, Karolinska Institutet, Stockholm, Sweden.

ABSTRACT

Background: The response of cells to TGFβ and EGF is mediated by a network of various intracellular regulators. The signaling crosstalk between different regulators is of key importance for tumorigenesis. The crosstalk may explain the modulation of cellular responses to the same regulator by another signaling molecule. As PKN1 - a serine/threonine kinase implicated in tumorigenesis - was identified as potential crosstalk node for TGFβ and EGF signaling, the cellular functions that may be affected by PKN1 in a crosstalk of TGFβ and EGF were explored.

Methods: To investigate the contribution of PKN1 to TGFβ and EGF signaling, transiently PKN1-transfected HEC-1-A endometrial cancer cells were generated and subjected to treatment with TGFβ1, EGF, and their combination. Proliferation, apoptosis, invasion, wound healing, and migration assays were performed. The impact of PKN1 on the expression and phosphorylation of intracellular proteins was monitored by immunoblotting.

Results: It was demonstrated that PKN1 modulated the responses of HEC-A-1 endometrial cancer cells to TGFβ1 and EGF. PKN1 had an inhibitory effect on the stimulation of cell migration, and PKN1 kinase activity was required for the inhibitory effect of TGFβ and EGF on cell proliferation and invasiveness. It was observed that phosphorylation of Smad2, FAK, and Erk1/2 correlated with responses of the cells to TGFβ1 and EGF.

Conclusion: PKN1 modulates TGFβ- and EGF-dependent regulation of cell proliferation, migration, and invasiveness, and therefore is a component of the network signaling downstream of TGFβ and EGF.

No MeSH data available.


Related in: MedlinePlus

PKN1 suppressed migration of cells stimulated by TGFβ1 and EGF.Notes: Columns show the total area of the membranes occupied by the cells which migrated through the membrane for HEC-1-A/PCDNA3, HEC-1-A/WT PKN1, HEC-1-A/KN PKN1, and HEC-1-A/CA PKN1 cells treated with TGFβ1, EGF, and their combination. Images show examples of membranes with stained cells. #Represents statistical significance (P<0.05) of observed differences among unpaired groups; *represents statistical significance (P<0.05) of observed differences among multiple groups. A representative experiment out of three is shown.Abbreviations: CA, constitutively active; KN, kinase negative; WT, wild type.
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f5-ott-7-1397: PKN1 suppressed migration of cells stimulated by TGFβ1 and EGF.Notes: Columns show the total area of the membranes occupied by the cells which migrated through the membrane for HEC-1-A/PCDNA3, HEC-1-A/WT PKN1, HEC-1-A/KN PKN1, and HEC-1-A/CA PKN1 cells treated with TGFβ1, EGF, and their combination. Images show examples of membranes with stained cells. #Represents statistical significance (P<0.05) of observed differences among unpaired groups; *represents statistical significance (P<0.05) of observed differences among multiple groups. A representative experiment out of three is shown.Abbreviations: CA, constitutively active; KN, kinase negative; WT, wild type.

Mentions: As the systemic analysis indicated that PKN1 may affect migration of cells, wound healing18 and membrane migration19 assays were performed (Figures 4 and 5). Wound healing assays explore migration capacities of cells that are under contact inhibition of proliferation. Membrane migration assays explore proliferating cells in a sparse culture. The molecular mechanisms triggering cell migration in these two tests may differ due to unequal conditions of tested cells. Despite differences, the tests may allow complementation of assessment of migration. The wound healing assay showed that PKN1 constructs prevented TGFβ1-induced closure (Figure 4). However, expression of the PKN1 constructs strongly promoted wound closure when cells were treated with both TGFβ1 and EGF (Figure 4).


PKN1 modulates TGFβ and EGF signaling in HEC-1-A endometrial cancer cell line.

Attarha S, Saini RK, Andersson S, Mints M, Souchelnytskyi S - Onco Targets Ther (2014)

PKN1 suppressed migration of cells stimulated by TGFβ1 and EGF.Notes: Columns show the total area of the membranes occupied by the cells which migrated through the membrane for HEC-1-A/PCDNA3, HEC-1-A/WT PKN1, HEC-1-A/KN PKN1, and HEC-1-A/CA PKN1 cells treated with TGFβ1, EGF, and their combination. Images show examples of membranes with stained cells. #Represents statistical significance (P<0.05) of observed differences among unpaired groups; *represents statistical significance (P<0.05) of observed differences among multiple groups. A representative experiment out of three is shown.Abbreviations: CA, constitutively active; KN, kinase negative; WT, wild type.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4128851&req=5

f5-ott-7-1397: PKN1 suppressed migration of cells stimulated by TGFβ1 and EGF.Notes: Columns show the total area of the membranes occupied by the cells which migrated through the membrane for HEC-1-A/PCDNA3, HEC-1-A/WT PKN1, HEC-1-A/KN PKN1, and HEC-1-A/CA PKN1 cells treated with TGFβ1, EGF, and their combination. Images show examples of membranes with stained cells. #Represents statistical significance (P<0.05) of observed differences among unpaired groups; *represents statistical significance (P<0.05) of observed differences among multiple groups. A representative experiment out of three is shown.Abbreviations: CA, constitutively active; KN, kinase negative; WT, wild type.
Mentions: As the systemic analysis indicated that PKN1 may affect migration of cells, wound healing18 and membrane migration19 assays were performed (Figures 4 and 5). Wound healing assays explore migration capacities of cells that are under contact inhibition of proliferation. Membrane migration assays explore proliferating cells in a sparse culture. The molecular mechanisms triggering cell migration in these two tests may differ due to unequal conditions of tested cells. Despite differences, the tests may allow complementation of assessment of migration. The wound healing assay showed that PKN1 constructs prevented TGFβ1-induced closure (Figure 4). However, expression of the PKN1 constructs strongly promoted wound closure when cells were treated with both TGFβ1 and EGF (Figure 4).

Bottom Line: The response of cells to TGFβ and EGF is mediated by a network of various intracellular regulators.It was observed that phosphorylation of Smad2, FAK, and Erk1/2 correlated with responses of the cells to TGFβ1 and EGF.PKN1 modulates TGFβ- and EGF-dependent regulation of cell proliferation, migration, and invasiveness, and therefore is a component of the network signaling downstream of TGFβ and EGF.

View Article: PubMed Central - PubMed

Affiliation: Department of Oncology-Pathology, Karolinska Institutet, Stockholm, Sweden ; Department of Women's and Children's Health, Karolinska Institutet, Stockholm, Sweden.

ABSTRACT

Background: The response of cells to TGFβ and EGF is mediated by a network of various intracellular regulators. The signaling crosstalk between different regulators is of key importance for tumorigenesis. The crosstalk may explain the modulation of cellular responses to the same regulator by another signaling molecule. As PKN1 - a serine/threonine kinase implicated in tumorigenesis - was identified as potential crosstalk node for TGFβ and EGF signaling, the cellular functions that may be affected by PKN1 in a crosstalk of TGFβ and EGF were explored.

Methods: To investigate the contribution of PKN1 to TGFβ and EGF signaling, transiently PKN1-transfected HEC-1-A endometrial cancer cells were generated and subjected to treatment with TGFβ1, EGF, and their combination. Proliferation, apoptosis, invasion, wound healing, and migration assays were performed. The impact of PKN1 on the expression and phosphorylation of intracellular proteins was monitored by immunoblotting.

Results: It was demonstrated that PKN1 modulated the responses of HEC-A-1 endometrial cancer cells to TGFβ1 and EGF. PKN1 had an inhibitory effect on the stimulation of cell migration, and PKN1 kinase activity was required for the inhibitory effect of TGFβ and EGF on cell proliferation and invasiveness. It was observed that phosphorylation of Smad2, FAK, and Erk1/2 correlated with responses of the cells to TGFβ1 and EGF.

Conclusion: PKN1 modulates TGFβ- and EGF-dependent regulation of cell proliferation, migration, and invasiveness, and therefore is a component of the network signaling downstream of TGFβ and EGF.

No MeSH data available.


Related in: MedlinePlus