Dlg-1 Interacts With and Regulates the Activities of Fibroblast Growth Factor Receptors and EphA2 in the Mouse Lens

Purpose We previously showed that Discs large-1 (Dlg-1) regulates lens fiber cell structure and the fibroblast growth factor receptor (Fgfr) signaling pathway, a pathway required for fiber cell differentiation. Herein, we investigated the mechanism through which Dlg-1 regulates Fgfr signaling. Methods Immunofluorescence was used to measure levels of Fgfr1, Fgfr2, and activated Fgfr signaling intermediates, pErk and pAkt, in control and Dlg-1–deficient lenses that were haplodeficient for Fgfr1 or Fgfr2. Immunoblotting was used to measure levels of N-cadherin, EphA2, β-catenin, and tyrosine-phosphorylated EphA2, Fgfr1, Fgfr2, and Fgfr3 in cytoskeletal-associated and cytosolic fractions of control and Dlg-1–deficient lenses. Complex formation between Dlg-1, N-cadherin, β-catenin, Fgfr1, Fgfr2, Fgfr3, and EphA2 was assessed by coimmunoprecipitation. Results Lenses deficient for Dlg-1 and haplodeficient for Fgfr1 or Fgfr2 showed increased levels of Fgfr2 or Fgfr1, respectively. Levels of pErk and pAkt correlated with the level of Fgfr2. N-cadherin was reduced in the cytoskeletal-associated fraction and increased in the cytosolic fraction of Dlg-1–deficient lenses. Dlg-1 complexed with β-catenin, EphA2, Fgfr1, Fgfr2, and Fgfr3. EphA2 complexed with N-cadherin, β-catenin, Fgfr1, Fgfr2, and Fgfr3. Levels of these interactions were altered in Dlg-1–deficient lenses. Loss of Dlg-1 led to changes in Fgfr1, Fgfr2, Fgfr3, and EphA2 levels and to greater changes in the levels of their activation. Conclusions Dlg-1 complexes with and regulates the activities of EphA2, Fgfr1, Fgfr2, and Fgfr3. As EphA2 contains a Psd95/Dlg/ZO-1 (PDZ) binding motif, whereas Fgfrs do not, we propose that the PDZ protein, Dlg-1, modulates Fgfr signaling through regulation of EphA2.

[1]  P. Tsonis,et al.  Comparative transcriptome analysis of epithelial and fiber cells in newborn mouse lenses with RNA sequencing , 2014, Molecular vision.

[2]  Sungkyoung Lee,et al.  Loss of Dlg-1 in the Mouse Lens Impairs Fibroblast Growth Factor Receptor Signaling , 2014, PloS one.

[3]  Jonathan A. Cooper,et al.  EphA2 and Src regulate equatorial cell morphogenesis during lens development , 2013, Development.

[4]  L. Walch.  Emerging Role of the Scaffolding Protein Dlg1 in Vesicle Trafficking , 2013, Traffic.

[5]  P. Lambert,et al.  Requirement for Dlgh-1 in Planar Cell Polarity and Skeletogenesis during Vertebrate Development , 2013, PloS one.

[6]  L. Reneker,et al.  Frs2α enhances fibroblast growth factor-mediated survival and differentiation in lens development , 2012, Development.

[7]  L. Banks,et al.  Regulation of the DLG tumor suppressor by β‐catenin , 2012, International journal of cancer.

[8]  Alexander I. Son,et al.  Human Cataract Mutations in EPHA2 SAM Domain Alter Receptor Stability and Function , 2012, PloS one.

[9]  S. Bassnett,et al.  A role for epha2 in cell migration and refractive organization of the ocular lens. , 2012, Investigative ophthalmology & visual science.

[10]  Catherine Cheng,et al.  Diverse Roles of Eph/ephrin Signaling in the Mouse Lens , 2011, PloS one.

[11]  V. Krishnappa,et al.  Fibroblast Growth Factor 2 (Fgf2) Inhibits Differentiation of Mesenchymal Stem Cells by Inducing Twist2 and Spry4, Blocking Extracellular Regulated Kinase Activation, and Altering Fgf Receptor Expression Levels , 2011, Stem cells.

[12]  F. Lovicu,et al.  Planar cell polarity in the mammalian eye lens , 2011, Organogenesis.

[13]  F. Lovicu,et al.  Understanding the role of growth factors in embryonic development: insights from the lens , 2011, Philosophical Transactions of the Royal Society B: Biological Sciences.

[14]  Y. Liu,et al.  The function of FGF signaling in the lens placode. , 2011, Developmental biology.

[15]  F. Lovicu,et al.  Growth factor signaling in vitreous humor-induced lens fiber differentiation. , 2010, Investigative ophthalmology & visual science.

[16]  K. Akimoto,et al.  A cell polarity protein aPKCλ is required for eye lens formation and growth , 2009, Developmental biology.

[17]  S. Shatadal,et al.  Cell‐autonomous requirements for Dlg‐1 for lens epithelial cell structure and fiber cell morphogenesis , 2009, Developmental dynamics : an official publication of the American Association of Anatomists.

[18]  T. Spector,et al.  EPHA2 Is Associated with Age-Related Cortical Cataract in Mice and Humans , 2009, PLoS genetics.

[19]  Alexander I. Son,et al.  Loss of ephrin-A5 function disrupts lens fiber cell packing and leads to cataract , 2008, Proceedings of the National Academy of Sciences.

[20]  T. Itakura,et al.  EphA4 promotes cell proliferation and migration through a novel EphA4-FGFR1 signaling pathway in the human glioma U251 cell line , 2008, Molecular Cancer Therapeutics.

[21]  M. Robinson,et al.  Fibroblast growth factor receptor signaling is essential for lens fiber cell differentiation. , 2008, Developmental biology.

[22]  C. Hudson,et al.  Ephrin-Eph signalling drives the asymmetric division of notochord/neural precursors in Ciona embryos , 2007, Development.

[23]  M. Robinson.  An essential role for FGF receptor signaling in lens development. , 2006, Seminars in cell & developmental biology.

[24]  L. Banks,et al.  Human discs large and scrib are localized at the same regions in colon mucosa and changes in their expression patterns are correlated with loss of tissue architecture during malignant progression , 2006, International journal of cancer.

[25]  J. Rossant,et al.  Fibroblast growth factor receptor 1 (Fgfr1) is not essential for lens fiber differentiation in mice. , 2006, Molecular vision.

[26]  A. Griep,et al.  Localization of PDZ domain containing proteins Discs Large-1 and Scribble in the mouse eye. , 2005, Molecular vision.

[27]  J. Schlessinger,et al.  Trans-activation of EphA4 and FGF receptors mediated by direct interactions between their cytoplasmic domains. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[28]  A. Bernstein,et al.  Mutagenesis of the epithelial polarity gene, discs large 1, perturbs nephrogenesis in the developing mouse kidney. , 2005, Kidney international.

[29]  R. Ashery-Padan,et al.  Signaling through FGF receptor‐2 is required for lens cell survival and for withdrawal from the cell cycle during lens fiber cell differentiation , 2005, Developmental dynamics : an official publication of the American Association of Anatomists.

[30]  L. Banks,et al.  Differential expression of the human homologue of drosophila discs large oncosuppressor in histologic samples from human papillomavirus–associated lesions as a marker for progression to malignancy , 2004, International journal of cancer.

[31]  P. Overbeek,et al.  Insertion of a Pax6 consensus binding site into the alphaA-crystallin promoter acts as a lens epithelial cell enhancer in transgenic mice. , 2004, Investigative ophthalmology & visual science.

[32]  S. Moody.  To Differentiate or Not to Differentiate: Regulation of Cell Fate Decisions by Being in the Right Place at the Right Time , 2004, Cell cycle.

[33]  Minh N. H. Nguyen,et al.  Requirement of PDZ-Containing Proteins for Cell Cycle Regulation and Differentiation in the Mouse Lens Epithelium , 2003, Molecular and Cellular Biology.

[34]  M. Ittmann,et al.  Inducible prostate intraepithelial neoplasia with reversible hyperplasia in conditional FGFR1-expressing mice. , 2003, Cancer research.

[35]  Mustafa Ozen,et al.  Conditional activation of fibroblast growth factor receptor (FGFR) 1, but not FGFR2, in prostate cancer cells leads to increased osteopontin induction, extracellular signal-regulated kinase activation, and in vivo proliferation. , 2003, Cancer research.

[36]  E. Olson,et al.  Conditional inactivation of FGF receptor 2 reveals an essential role for FGF signaling in the regulation of osteoblast function and bone growth , 2003, Development.

[37]  M. Feldmeier,et al.  Evaluation of fibroblast growth factor signaling during lens fiber cell differentiation. , 2003, Investigative ophthalmology & visual science.

[38]  C. Deng,et al.  Generation of Fgfr1 conditional knockout mice , 2002, Genesis.

[39]  W. Lim,et al.  Mechanism and role of PDZ domains in signaling complex assembly. , 2001, Journal of cell science.

[40]  I. Daar,et al.  Fibroblast Growth Factor Receptor-Mediated Rescue of x-Ephrin B1-Induced Cell Dissociation in XenopusEmbryos , 2000, Molecular and Cellular Biology.

[41]  E. Pasquale,et al.  Signal transfer by Eph receptors , 1999, Cell and Tissue Research.

[42]  I. Daar,et al.  Fibroblast Growth Factor Receptor-Mediated Rescue of x-Ephrin B1-Induced Cell Dissociation in Xenopus Embryos , 1999 .

[43]  F. Lovicu,et al.  Lens development , 1999, Eye.

[44]  R. Huganir,et al.  PDZ Proteins Bind, Cluster, and Synaptically Colocalize with Eph Receptors and Their Ephrin Ligands , 1998, Neuron.

[45]  R. D. Iongh,et al.  Differential expression of fibroblast growth factor receptors during rat lens morphogenesis and growth. , 1997, Investigative ophthalmology & visual science.

[46]  J. Liu,et al.  IGF enhancement of FGF-induced fibre differentiation and DNA synthesis in lens explants. , 1996, Experimental eye research.

[47]  X. Gu,et al.  Fibroblast growth factor 2 uses distinct signaling pathways for cell proliferation and cell shape changes in corneal endothelial cells. , 1996, Investigative ophthalmology & visual science.

[48]  Y. Benjamini,et al.  Controlling the false discovery rate: a practical and powerful approach to multiple testing , 1995 .

[49]  J. McAvoy,et al.  IGF-1 enhancement of FGF-induced lens fiber differentiation in rats of different ages. , 1993, Investigative ophthalmology & visual science.

[50]  J. Piatigorsky,et al.  Lens differentiation in vertebrates. A review of cellular and molecular features. , 1981, Differentiation; research in biological diversity.