Hrg induces TGFb2 expression and SMAD3 phosphorylation in breast cancer cells

LARA ANGELA; KORDON EDITH; GONZALEZ AGUSTIN; DE LAURENTIIS ANDREA; ABBA MARTIN; WERTHEIMER, EVA V.

Bariloche

Simposio; Fourth South American Spring Symposium in Signal Transduction and Molecular Medicine? (SISTAM 2018); 2018

IUBMB focused meeting

Rac1 is a Rho-GTPase that promotes migration of breast cancer cells. Heregulin (HRG) activates Rac1 in T-47D breast cancer cells through the tyrosine kinase receptor, ErbB3, in a time-dependent manner. Rac1 activation increases rapidly with a peak at 5 min and is maintained for at least 1 h, followed by a gradual decrease at 6 h post-stimulation with HRG. Previous data indicate that Rac1 activation by HRG leads to an increase in TGFb2 mRNA levels, with a significant increase after 30 min and a maximum of expression at 2 h. In addition, wound healing assays revealed that TGFb2 is able to induce cell migration of breast cancer cells in a dose-dependent manner. However, the combined addition of HRG and TGFb2 produced the same increase in cell migration as HRG alone. These results suggest that Rac1 activation peak precedes the maximal increase of TGFβ2. Therefore, we propose that this cytokine is necessary for the maintenance of HRG-triggered phenotype of breast cancer cells. The aim of this work is to study the effect of ErbB3 activation by HRG on members of the TGFb2 signaling cascade and the possibility of crosstalk between both pathways. First, we observed that TGFb2 promotes phosphorylation of Smad3, a downstream effector of TGFB receptors. However, TGFb2 is not able to induce Rac1 activation by itself neither at 5 nor 60 min after stimulus addition. We determined that HRG induces Smad3 phosphorylation in a time-dependent manner that follows the same kinetics of Rac1 activation by this same ligand. HRG-induced Smad3 phosphorylation is not affected by H89 (a PKA inhibitor) but it is inhibited by OU126 (an Erk inhibitor). In addition, HRG induces an increase of TGFBR1, TGFBR2, and SMAD3 mRNA levels while it downregulates SMAD4 expression. Our results suggest that crosstalk between ErbB and TGFb signaling cascades is necessary for HRG-triggered cell migration of breast cancer cells.