title: Wnt activation by wild type and mutant myocilin in cultured human trabecular meshwork cells.
abstract: BACKGROUND
  Myocilin is a gene linked to the most prevalent form of glaucoma, a major blinding disease. The trabecular meshwork (TM), a specialized eye tissue, is believed to be involved, at least in part, in the development of glaucoma. The Pro³⁷⁰ to Leu (P370L) mutation of myocilin is associated with severe glaucoma phenotypes and Gln³⁶⁸ stop (Q368X) is the most common myocilin mutation reported. Myocilin, upon overexpression, has been shown to induce phenotypes that include a loss of actin stress fibers, an increase in the cAMP level and protein kinase A (PKA) activity, as well as a reduction in the RhoA activity. We examined herein whether Wnt signaling pathway is involved in the myocilin phenotypes and whether P370L and Q368X mutants also display biological effects similar to those of the wild type myocilin.
  
  
  RESULTS
  Wild type myocilin, when transfected into cultured human TM cells, induced a loss of actin stress fibers as judged by phalloidin staining. Such a loss was averted by treatment of secreted Frizzled-related protein 1 (sFRP1), an inhibitor of Wnt signaling. Consistent with the notion that Wnt pathway mediates the myocilin phenotype, Wnt activation was demonstrated by TOP/FOP-Flash reporter assays. Treatment of human TM cells of a Wnt activator, SB216763, as well as transfection of myocilin P370L and Q368X mutants all resulted in actin stress fiber loss, PKA activation and RhoA inactivation. The PKA elevation was obviated by the sFRP1 treatment, indicating that Wnt signaling was upstream that of PKA.
  
  
  CONCLUSIONS
  The present study demonstrated that following forced expression of wild type myocilin, Wnt was activated, triggering in turn other myocilin-related alterations. P370L and Q368X mutations induced similar phenotypes, suggesting one possible mechanism how the mutants may lead to TM cell damage and pathology.
authors.affiliation: Department of Ophthalmology and Visual Sciences, University of Illinois at Chicago College of Medicine, Chicago, Illinois, United States of America.
authors.forename: Xiang
authors.initials: X
authors.lastname: Shen
authors.forename: Hongyu
authors.initials: H
authors.lastname: Ying
authors.forename: Beatrice Y J T
authors.initials: BY
authors.lastname: Yue
mesh_terms: D000199:Actins / Q000378:metabolism; D000328:Adult; D018938:Caco-2 Cells; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000235:genetics* / Q000378:metabolism*; D005136:Eye Proteins / Q000235:genetics* / Q000378:metabolism*; D022001:Focal Adhesions / Q000187:drug effects / Q000378:metabolism; D006023:Glycoproteins / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins; D008301:Maleimides / Q000494:pharmacology; D008875:Middle Aged; D009154:Mutation; D010641:Phenotype; D011506:Proteins / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects; D014129:Trabecular Meshwork / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D014162:Transfection; D051153:Wnt Proteins / Q000378:metabolism*; D055815:Young Adult; D000099179:Myocilin; D000099274:Frizzled-Related Proteins
chemical_list: D000199:Actins; D017868:Cyclic AMP-Dependent Protein Kinases; D003598:Cytoskeletal Proteins; D005136:Eye Proteins; D006023:Glycoproteins; D007211:Indoles; D047908:Intracellular Signaling Peptides and Proteins; D008301:Maleimides; D011506:Proteins; D051153:Wnt Proteins; D000099179:Myocilin; D000099274:Frizzled-Related Proteins; C417521:SB 216763