title: Transmembrane/cytoplasmic, rather than catalytic, domains of Mmp14 signal to MAPK activation and mammary branching morphogenesis via binding to integrin β1.
abstract: Epithelial cell invasion through the extracellular matrix (ECM) is a crucial step in branching morphogenesis. The mechanisms by which the mammary epithelium integrates cues from the ECM with intracellular signaling in order to coordinate invasion through the stroma to make the mammary tree are poorly understood. Because the cell membrane-bound matrix metalloproteinase Mmp14 is known to play a key role in cancer cell invasion, we hypothesized that it could also be centrally involved in integrating signals for mammary epithelial cells (MECs) to navigate the collagen 1 (CL-1)-rich stroma of the mammary gland. Expression studies in nulliparous mice that carry a NLS-lacZ transgene downstream of the Mmp14 promoter revealed that Mmp14 is expressed in MECs at the tips of the branches. Using both mammary organoids and 3D organotypic cultures, we show that MMP activity is necessary for invasion through dense CL-1 (3 mg/ml) gels, but dispensable for MEC branching in sparse CL-1 (1 mg/ml) gels. Surprisingly, however, Mmp14 without its catalytic activity was still necessary for branching. Silencing Mmp14 prevented cell invasion through CL-1 and disrupted branching altogether; it also reduced integrin β1 (Itgb1) levels and attenuated MAPK signaling, disrupting Itgb1-dependent invasion/branching within CL-1 gels. FRET imaging revealed that Mmp14 associates directly with Itgb1. We identified a domain of Mmp14 that is required for modulating the levels of Itgb1, MEC signaling and the rate of invasion within CL-1. These results shed light on hitherto undescribed non-proteolytic activities of Mmp14 that are necessary for the Itgb1-dependent biochemical and mechanical signals that regulate branching in the mammary epithelium.
authors.affiliation: Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA. hmori@lbl.gov
authors.forename: Hidetoshi
authors.initials: H
authors.lastname: Mori
authors.forename: Alvin T
authors.initials: AT
authors.lastname: Lo
authors.forename: Jamie L
authors.initials: JL
authors.lastname: Inman
authors.forename: Jordi
authors.initials: J
authors.lastname: Alcaraz
authors.forename: Cyrus M
authors.initials: CM
authors.lastname: Ghajar
authors.forename: Joni D
authors.initials: JD
authors.lastname: Mott
authors.forename: Celeste M
authors.initials: CM
authors.lastname: Nelson
authors.forename: Connie S
authors.initials: CS
authors.lastname: Chen
authors.forename: Hui
authors.initials: H
authors.lastname: Zhang
authors.forename: Jamie L
authors.initials: JL
authors.lastname: Bascom
authors.forename: Motoharu
authors.initials: M
authors.lastname: Seiki
authors.forename: Mina J
authors.initials: MJ
authors.lastname: Bissell
mesh_terms: D000818:Animals; D020134:Catalytic Domain; D002462:Cell Membrane / Q000378:metabolism*; D003094:Collagen / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods; D020868:Gene Silencing; D019012:Integrin beta1 / Q000378:metabolism*; D016086:Lentivirus / Q000378:metabolism; D020935:MAP Kinase Signaling System; D008321:Mammary Glands, Animal / Q000378:metabolism; D053511:Matrix Metalloproteinase 14 / Q000378:metabolism*; D051379:Mice; D008822:Mice, Transgenic; D008954:Models, Biological; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D015398:Signal Transduction; D019076:Transgenes
chemical_list: D019012:Integrin beta1; C507096:Mmp14 protein, mouse; D003094:Collagen; D053511:Matrix Metalloproteinase 14