title: RhoA is down-regulated at cell-cell contacts via p190RhoGAP-B in response to tensional homeostasis.
abstract: Breast epithelial cells cultured in three-dimensional (3D) collagen gels undergo ductal morphogenesis when the gel is compliant and they can achieve tensional homeostasis. We previously showed that this process requires down-regulation of Rho in compliant collagen gels, but the mechanism remains undefined. In this study, we find that p190RhoGAP-B, but not p190RhoGAP-A, mediates down-regulation of RhoA activity and ductal morphogenesis in T47D cells cultured in compliant 3D collagen gels. In addition, both RhoA and p190RhoGAP-B colocalize with p120-catenin at sites of cell-cell contact. The association between p190RhoGAP-B and p120-catenin is regulated by matrix compliance such that it increases in compliant vs. rigid collagen gels. Furthermore, knockdown of p120-catenin disrupts ductal morphogenesis, disregulates RhoA activity, and results in loss of p190B at cell-cell contacts. Consistent with these findings, using a RhoA-specific FRET biosensor (RhoA-FLARE.sc), we determined spatial RhoA activity to be significantly decreased at cell-cell contacts versus cell-ECM adhesions, and, of importance, spatial RhoA activity is regulated by p190B. This finding suggests that RhoA exists as an inactive pool at cell-cell contacts and is recruited to cell-ECM contacts within stiff matrices. Overall, these results demonstrate that RhoA is down-regulated at cell-cell contacts through p190RhoGAP-B, which is localized to cell-cell contacts by association with p120-catenin that is regulated by tensional homeostasis.
authors.affiliation: Department of Cellular and Regenerative Biology, University of Wisconsin-Madison, Madison, WI 53706, USA. ponik@wisc.edu
authors.forename: Suzanne M
authors.initials: SM
authors.lastname: Ponik
authors.forename: Steven M
authors.initials: SM
authors.lastname: Trier
authors.forename: Michele A
authors.initials: MA
authors.lastname: Wozniak
authors.forename: Kevin W
authors.initials: KW
authors.lastname: Eliceiri
authors.forename: Patricia J
authors.initials: PJ
authors.lastname: Keely
mesh_terms: D051177:Catenins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D002448:Cell Adhesion; D002450:Cell Communication; D018929:Cell Culture Techniques; D002460:Cell Line; D003094:Collagen / Q000737:chemistry; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism*; D005260:Female; D036681:Fluorescence Recovery After Photobleaching; D020690:GTPase-Activating Proteins / Q000235:genetics* / Q000378:metabolism; D005782:Gels; D005786:Gene Expression Regulation; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D007365:Intercellular Junctions / Q000378:metabolism* / Q000648:ultrastructure; D042361:Mammary Glands, Human / Q000166:cytology / Q000378:metabolism*; D018583:Pliability; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction; D013500:Surface Tension
chemical_list: C074560:ARHGAP35 protein, human; C513043:ARHGAP5 protein, human; D051177:Catenins; D020690:GTPase-Activating Proteins; D005782:Gels; D020662:Guanine Nucleotide Exchange Factors; D012097:Repressor Proteins; D003094:Collagen
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