P190B RhoGAP Overexpression in the Developing Mammary Epithelium Induces TGFβ-dependent Fibroblast Activation

Melissa Gillette, Kristi Bray, Alisa Blumenthaler, Tracy Vargo-Gogola

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

Rho GTPases mediate stromal-epithelial interactions that are important for mammary epithelial cell (MEC) morphogenesis. Increased extracellular matrix (ECM) deposition and reorganization affect MEC morphogenesis in a Rho GTPase-dependent manner. Although the effects of altered ECM on MEC morphogenesis have been described, how MECs regulate stromal deposition is not well understood. Previously, we showed that p190B RhoGAP overexpression disrupts mammary gland morphogenesis by inducing hyperbranching in association with stromal alterations. We therefore hypothesized that MEC overexpression of p190B regulates paracrine interactions to impact fibroblast activation. Using a combination of in vivo morphometric and immunohistochemical analyses and primary cell culture assays, we found that p190B overexpression in MECs activates fibroblasts leading to increased collagen, fibronectin, and laminin production and elevated expression of the collagen crosslinking enzyme lysyl oxidase. Phosphorylation of the TGF-β effector SMAD2 and expression of the TGF-β target gene αSma were increased in p190B-associated fibroblasts, suggesting that elevated TGF-β signaling promoted fibroblast activation. Mechanical tension and TGF-β cooperate to activate fibroblasts. Interestingly, active TGF-β was elevated in conditioned medium from p190B overexpressing MECs compared to control MECs, and p190B overexpressing MECs exhibited increased contractility in a collagen gel contraction assay. These data suggest that paracrine signaling from the p190B overexpressing MECs may activate TGF-β signaling in adjacent fibroblasts. In support of this, transfer of conditioned medium from p190B overexpressing MECs onto wildtype fibroblasts or co-culture of p190B overexpressing MECs with wildtype fibroblasts increased SMAD2 phosphorylation and mRNA expression of ECM genes in the fibroblasts when compared to fibroblasts treated with control CM or co-cultured with control MECs. The increased ECM gene expression and SMAD2 phosphorylation were blocked by treatment with a TGF-β receptor inhibitor. Taken together, these data suggest that p190B overexpression in the mammary epithelium induces fibroblast activation via elevated TGF-β paracrine signaling.

Original languageEnglish (US)
Article numbere65105
JournalPLoS One
Volume8
Issue number5
DOIs
StatePublished - May 22 2013

Fingerprint

Fibroblasts
breasts
fibroblasts
Breast
epithelium
Epithelium
Chemical activation
Morphogenesis
extracellular matrix
Phosphorylation
morphogenesis
Extracellular Matrix
epithelial cells
Epithelial Cells
Paracrine Communication
collagen
phosphorylation
rho GTP-Binding Proteins
Collagen
guanosinetriphosphatase

ASJC Scopus subject areas

  • Agricultural and Biological Sciences(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Medicine(all)

Cite this

P190B RhoGAP Overexpression in the Developing Mammary Epithelium Induces TGFβ-dependent Fibroblast Activation. / Gillette, Melissa; Bray, Kristi; Blumenthaler, Alisa; Vargo-Gogola, Tracy.

In: PLoS One, Vol. 8, No. 5, e65105, 22.05.2013.

Research output: Contribution to journalArticle

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