title: A syndecan-4 binding peptide derived from laminin 5 uses a novel PKCε pathway to induce cross-linked actin network (CLAN) formation in human trabecular meshwork (HTM) cells.
abstract: In this study, we examined the role(s) of syndecan-4 in regulating the formation of an actin geodesic dome structure called a cross-linked actin network (CLAN) in which syndecan-4 has previously been localized. CLANs have been described in several different cell types, but they have been most widely studied in human trabecular meshwork (HTM) cells where they may play a key role in controlling intraocular pressure by regulating aqueous humor outflow from the eye. In this study we show that a loss of cell surface synedcan-4 significantly reduces CLAN formation in HTM cells. Analysis of HTM cultures treated with or without dexamethasone shows that laminin 5 deposition within the extracellular matrix is increased by glucocorticoid treatment and that a laminin 5-derived, syndecan-4-binding peptide (PEP75), induces CLAN formation in TM cells. This PEP75-induced CLAN formation was inhibited by heparin and the broad spectrum PKC inhibitor Ro-31-7549. In contrast, the more specific PKCα inhibitor Gö 6976 had no effect, thus excluding PKCα as a downstream effector of syndecan-4 signaling. Analysis of PKC isozyme expression showed that HTM cells also expressed both PKCγ and PKCε. Cells treated with a PKCε agonist formed CLANs while a PKCα/γ agonist had no effect. These data suggest that syndecan-4 is essential for CLAN formation in HTM cells and that a novel PKCε-mediated signaling pathway can regulate formation of this unique actin structure.
authors.affiliation: Department of Ophthalmology & Visual Sciences, Medical Science Center, 1300 University Avenue, Madison, WI 53706, United States. Electronic address: msfilla@wisc.edu.
authors.forename: Mark S
authors.initials: MS
authors.lastname: Filla
authors.affiliation: Department of Pathology & Laboratory Medicine, Medical Science Center, 1300 University Avenue, Madison, WI 53706, United States. Electronic address: rclark2@wisc.edu.
authors.forename: Ross
authors.initials: R
authors.lastname: Clark
authors.affiliation: Department of Ophthalmology & Visual Sciences, Medical Science Center, 1300 University Avenue, Madison, WI 53706, United States; Department of Pathology & Laboratory Medicine, Medical Science Center, 1300 University Avenue, Madison, WI 53706, United States. Electronic address: dmpeter2@wisc.edu.
authors.forename: Donna M
authors.initials: DM
authors.lastname: Peters
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism*; D000199:Actins / Q000235:genetics / Q000378:metabolism*; D000293:Adolescent; D000328:Adult; D000893:Anti-Inflammatory Agents / Q000494:pharmacology; D015153:Blotting, Western; D015815:Cell Adhesion Molecules / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D003432:Cross-Linking Reagents / Q000494:pharmacology; D003907:Dexamethasone / Q000494:pharmacology; D005128:Eye Diseases / Q000378:metabolism / Q000473:pathology; D005455:Fluorescent Antibody Technique; D006801:Humans; D010446:Peptide Fragments / Q000378:metabolism*; D051744:Protein Kinase C-epsilon / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D053671:Syndecan-4 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D014129:Trabecular Meshwork / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D000097794:Kalinin
chemical_list: D000199:Actins; D000893:Anti-Inflammatory Agents; D015815:Cell Adhesion Molecules; D003432:Cross-Linking Reagents; D003907:Dexamethasone; D010446:Peptide Fragments; D051744:Protein Kinase C-epsilon; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D053671:Syndecan-4; D000097794:Kalinin
keywords: CLANs; Cytoskeleton; Laminin 5; Protein kinase C; Syndecan-4; Trabecular meshwork