Signaling pathways activated by epidermal growth factor receptor or fibroblast growth factor receptor differentially regulate branching morphogenesis in fetal mouse submandibular glands

Although growth factor signaling is required for embryonic development of organs, individual signaling mechanisms regulating these organotypic processes are just beginning to be defined. We compared signaling activated in fetal mouse submandibular glands (SMGs) by three growth factors, epidermal growth factor (EGF), fibroblast growth factor (FGF) 7, or FGF10, and correlated it with specific events of branching morphogenesis. Immunoblotting showed that EGF strongly stimulated phosphorylation of extracellular signal‐regulated kinase‐1/2 (ERK‐1/2) and weakly stimulated phosphorylation of phospholipase Cγ1 (PLCγ1) and phosphatidylinositol‐3 kinase (PI3K) in cultured E14 SMG. However, FGF7 and FGF10 stimulated phosphorylation of both PLCγ1 and PI3K, but elicited only minimal phosphorylation of ERK‐1/2. Morphological study of mesenchyme‐free SMG epithelium cultured in Matrigel revealed that EGF induced cleft formation of endpieces, that FGF7 stimulated both cleft formation and stalk elongation, but that FGF10 induced only stalk elongation. In mesenchyme‐free SMG epithelium cultured with EGF, FGF7 and FGF10, U0126 (MEK inhibitor) completely blocked cleft formation, whereas U73122 (PLCγ1 inhibitor) suppressed stalk elongation. These finding suggest that EGF stimulates cleft formation and drives branch formation via ERK‐1/2, and that FGF7 stimulates both cleft formation and stalk elongation via PLCγ1 and partly via ERK‐1/2, but that FGF10 stimulates stalk elongation mainly via PLCγ1.

[1]  E. Borghese.  Explantation experiments on the influence of the connective tissue capsule on the development of the epithelial part of the submandibular gland of Mus musculus. , 1950, Journal of anatomy.

[2]  E. Borghese.  The development in vitro of the submandibular and sublingual glands of Mus musculus. , 1950, Journal of anatomy.

[3]  John Tyler Bonner,et al.  Morphogenesis , 1965, Cell.

[4]  B. Spooner,et al.  Extracellular matrix involvement in epithelial branching morphogenesis. , 1986, Developmental biology.

[5]  H Nogawa,et al.  Substitution for mesenchyme by basement-membrane-like substratum and epidermal growth factor in inducing branching morphogenesis of mouse salivary epithelium. , 1991, Development.

[6]  C. MacArthur,et al.  Receptor Specificity of the Fibroblast Growth Factor Family* , 1996, The Journal of Biological Chemistry.

[7]  M. Kashimata,et al.  Epidermal growth factor system is a physiological regulator of development of the mouse fetal submandibular gland and regulates expression of the α6‐integrin subunit , 1997 .

[8]  Y. Yarden,et al.  The ErbB signaling network in embryogenesis and oncogenesis: signal diversification through combinatorial ligand‐receptor interactions , 1997, FEBS letters.

[9]  G. Martin,et al.  The roles of FGFs in the early development of vertebrate limbs. , 1998, Genes & development.

[10]  H Nogawa,et al.  EGF‐dependent lobule formation and FGF7‐dependent stalk elongation in branching morphogenesis of mouse salivary epithelium in vitro , 1999, Developmental dynamics : an official publication of the American Association of Anatomists.

[11]  M. Melnick,et al.  Submandibular gland morphogenesis: stage-specific expression of TGF-alpha/EGF, IGF, TGF-beta, TNF, and IL-6 signal transduction in normal embryonic mice and the phenotypic effects of TGF-beta2, TGF-beta3, and EGF-r null mutations. , 1999, The Anatomical record.

[12]  M. Melnick,et al.  Submandibular gland morphogenesis: Stage‐specific expression of TGF‐α/EGF, IGF, TGF‐β, TNF, and IL‐6 signal transduction in normal embryonic mice and the phenotypic effects of TGF‐β2, TGF‐β3, and EGF‐r null mutations , 1999 .

[13]  B. Spencer‐Dene,et al.  An important role for the IIIb isoform of fibroblast growth factor receptor 2 (FGFR2) in mesenchymal-epithelial signalling during mouse organogenesis. , 2000, Development.

[14]  C. Powers,et al.  Fibroblast growth factors, their receptors and signaling. , 2000, Endocrine-related cancer.

[15]  M. Melnick,et al.  Mouse submandibular gland morphogenesis: a paradigm for embryonic signal processing. , 2000, Critical reviews in oral biology and medicine : an official publication of the American Association of Oral Biologists.

[16]  S. Kato,et al.  FGF10 acts as a major ligand for FGF receptor 2 IIIb in mouse multi-organ development. , 2000, Biochemical and biophysical research communications.

[17]  T. Faraggiana,et al.  The ERK-1/2 signaling pathway is involved in the stimulation of branching morphogenesis of fetal mouse submandibular glands by EGF. , 2000, Developmental biology.

[18]  Y. Yarden,et al.  Untangling the ErbB signalling network , 2001, Nature Reviews Molecular Cell Biology.

[19]  S. Higashiyama,et al.  Involvement of heparin-binding EGF-like growth factor and its processing by metalloproteinases in early epithelial morphogenesis of the submandibular gland. , 2001, Developmental biology.

[20]  P. Itäranta,et al.  Expression of Sprouty genes 1, 2 and 4 during mouse organogenesis , 2001, Mechanisms of Development.

[21]  Y. Chai,et al.  Embryonic Submandibular Gland Morphogenesis: Stage-Specific Protein Localization of FGFs, BMPs, Pax6 and Pax9 in Normal Mice and Abnormal SMG Phenotypes in FgfR2-IIIc+/Δ, BMP7–/– and Pax6–/–Mice , 2001, Cells Tissues Organs.

[22]  H. Kleinman,et al.  Gene expression profiles of mouse submandibular gland development: FGFR1 regulates branching morphogenesis in vitro through BMP- and FGF-dependent mechanisms , 2002, Development.

[23]  N. Koyama,et al.  EGF‐stimulated signaling by means of PI3K, PLCγ1, and PKC isozymes regulates branching morphogenesis of the fetal mouse submandibular gland , 2003, Developmental dynamics : an official publication of the American Association of Anatomists.

[24]  Antony W Burgess,et al.  Epidermal growth factor receptor: mechanisms of activation and signalling. , 2003, Experimental cell research.

[25]  M. Mohammadi,et al.  Structural basis by which alternative splicing confers specificity in fibroblast growth factor receptors , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[26]  N. Rosenblum,et al.  Genetic Regulation of Branching Morphogenesis: Lessons Learned from Loss-of-Function Phenotypes , 2003, Pediatric Research.

[27]  Melinda Larsen,et al.  Role of PI 3-kinase and PIP3 in submandibular gland branching morphogenesis. , 2003, Developmental biology.

[28]  J. Rossant,et al.  Spatial and temporal patterns of ERK signaling during mouse embryogenesis , 2003, Development.

[29]  Derrick E Rancourt,et al.  Induction of chondro-, osteo- and adipogenesis in embryonic stem cells by bone morphogenetic protein-2: Effect of cofactors on differentiating lineages , 2005, BMC Developmental Biology.

[30]  Joseph Schlessinger,et al.  Common and Distinct Elements in Cellular Signaling via EGF and FGF Receptors , 2004, Science.

[31]  P. Carlsson,et al.  Sonic hedgehog signaling plays an essential role during embryonic salivary gland epithelial branching morphogenesis , 2004, Developmental dynamics : an official publication of the American Association of Anatomists.

[32]  A. Moon,et al.  FGF8 dose-dependent regulation of embryonic submandibular salivary gland morphogenesis. , 2004, Developmental biology.

[33]  Wei Zhang,et al.  WNK1 Activates ERK5 by an MEKK2/3-dependent Mechanism* , 2004, Journal of Biological Chemistry.

[34]  Y. Miyazaki,et al.  Tissue interaction mediated by neuregulin‐1 and ErbB receptors regulates epithelial morphogenesis of mouse embryonic submandibular gland , 2004, Developmental dynamics : an official publication of the American Association of Anatomists.

[35]  D. Ambrosetti,et al.  Mechanisms underlying differential responses to FGF signaling. , 2005, Cytokine & growth factor reviews.

[36]  M. Melnick,et al.  Embryonic Salivary Gland Branching Morphogenesis , 2005 .

[37]  Melinda Larsen,et al.  FGFR2b signaling regulates ex vivo submandibular gland epithelial cell proliferation and branching morphogenesis , 2005, Development.

[38]  S. Yamashina,et al.  Salivary gland morphogenesis and basement membranes , 2005, Anatomical science international.

[39]  Michael Melnick,et al.  FGF10/FGFR2b signaling plays essential roles during in vivo embryonic submandibular salivary gland morphogenesis , 2005, BMC Developmental Biology.

[40]  Y. Yarden,et al.  Phosphorylation of Carboxyl-terminal Tyrosines Modulates the Specificity of Sprouty-2 Inhibition of Different Signaling Pathways* , 2005, Journal of Biological Chemistry.

[41]  X. Nie,et al.  FGF signalling in craniofacial development and developmental disorders. , 2006, Oral diseases.

[42]  A. Citri,et al.  EGF–ERBB signalling: towards the systems level , 2006, Nature Reviews Molecular Cell Biology.

[43]  Kenneth M. Yamada,et al.  The matrix reorganized: extracellular matrix remodeling and integrin signaling. , 2006, Current opinion in cell biology.

[44]  Melinda Larsen,et al.  Cell and fibronectin dynamics during branching morphogenesis , 2006, Journal of Cell Science.

[45]  M. Hoffman,et al.  Salivary gland branching morphogenesis. , 2006, Differentiation; research in biological diversity.

[46]  E. Georges-Labouesse,et al.  Laminin alpha5 is necessary for submandibular gland epithelial morphogenesis and influences FGFR expression through beta1 integrin signaling. , 2007, Developmental biology.

[47]  Jeffrey H. Miner,et al.  Laminin α5 is necessary for submandibular gland epithelial morphogenesis and influences FGFR expression through β1 integrin signaling , 2007 .

[48]  Michael Elkin,et al.  Heparanase cleavage of perlecan heparan sulfate modulates FGF10 activity during ex vivo submandibular gland branching morphogenesis , 2007, Development.

[49]  A. Tucker,et al.  Salivary gland development. , 2007, Seminars in cell & developmental biology.

[50]  Erez M. Bublil,et al.  The EGF receptor family : spearheading a merger of signaling and therapeutics , 2007 .

[51]  J. Fata,et al.  The MAPK(ERK-1,2) pathway integrates distinct and antagonistic signals from TGFalpha and FGF7 in morphogenesis of mouse mammary epithelium. , 2007, Developmental biology.

[52]  Eytan Domany,et al.  A module of negative feedback regulators defines growth factor signaling , 2007, Nature Genetics.

[53]  Ido Amit,et al.  Regulation of MAPKs by growth factors and receptor tyrosine kinases. , 2007, Biochimica et biophysica acta.