title: Mechanistic basis of Rho GTPase-induced extracellular matrix synthesis in trabecular meshwork cells.
abstract: Elevated intraocular pressure arising from impaired aqueous humor drainage through the trabecular pathway is a major risk factor for glaucoma. To understand the molecular basis for Rho GTPase-mediated resistance to aqueous humor drainage, we investigated the possible interrelationship between actomyosin contractile properties and extracellular matrix (ECM) synthesis in human trabecular meshwork (TM) cells expressing a constitutively active form of RhoA (RhoAV14). TM cells expressing RhoAV14 exhibited significant increases in fibronectin, tenascin C, laminin, alpha-smooth muscle actin (alpha-SMA) levels, and matrix assembly in association with increased actin stress fibers and myosin light-chain phosphorylation. RhoAV14-induced changes in ECM synthesis and actin cytoskeletal reorganization were mimicked by lysophosphatidic acid and TGF-beta(2), known to increase resistance to aqueous humor outflow and activate Rho/Rho kinase signaling. RhoAV14, lysophosphatidic acid, and TGF-beta(2) stimulated significant increases in Erk1/2 phosphorylation, paralleled by profound increases in fibronectin, serum response factor (SRF), and alpha-SMA expression. Treatment of RhoA-activated TM cells with inhibitors of Rho kinase or Erk, on the other hand, decreased fibronectin and alpha-SMA levels. Although suppression of SRF expression (both endogenous and RhoA, TGF-beta(2)-stimulated) via the use of short hairpin RNA decreased alpha-SMA levels, fibronectin was unaffected. Conversely, fibronectin induced alpha-SMA expression in an SRF-dependent manner. Collectively, data on RhoA-induced changes in actomyosin contractile activity, ECM synthesis/assembly, and Erk activation, along with fibronectin-induced alpha-SMA expression in TM cells, reveal a potential molecular interplay between actomyosin cytoskeletal tension and ECM synthesis/assembly. This interaction could be significant for the homeostasis of aqueous humor drainage through the pressure-sensitive trabecular pathway.
authors: Dept. of Ophthalmology, AERI Bldg., Rm. 4010, Duke Univ. School of Medicine, Durham, NC 27710, USA.
authors: Padmanabhan P
authors: PP
authors: Pattabiraman
authors: Ponugoti Vasantha
authors: PV
authors: Rao
mesh_terms: D000199:Actins / Q000378:metabolism; D000205:Actomyosin / Q000378:metabolism; D000256:Adenoviridae / Q000235:genetics; D001082:Aqueous Humor / Q000378:metabolism; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000378:metabolism*; D004789:Enzyme Activation; D005109:Extracellular Matrix / Q000378:metabolism*; D016326:Extracellular Matrix Proteins / Q000096:biosynthesis*; D005353:Fibronectins / Q000096:biosynthesis; D005822:Genetic Vectors; D006801:Humans; D007797:Laminin / Q000096:biosynthesis; D008246:Lysophospholipids / Q000378:metabolism; D040542:Mechanotransduction, Cellular; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D018994:Myosin Light Chains / Q000378:metabolism; D010766:Phosphorylation; D034622:RNA Interference; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D026362:Serum Response Factor / Q000235:genetics / Q000378:metabolism; D022502:Stress Fibers / Q000378:metabolism; D019063:Tenascin / Q000096:biosynthesis; D014129:Trabecular Meshwork / Q000166:cytology / Q000201:enzymology*; D014161:Transduction, Genetic; D053781:Transforming Growth Factor beta2 / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*
chemical_list: D000199:Actins; D016326:Extracellular Matrix Proteins; D005353:Fibronectins; D007797:Laminin; D008246:Lysophospholipids; D018994:Myosin Light Chains; D011993:Recombinant Fusion Proteins; D026362:Serum Response Factor; D019063:Tenascin; D053781:Transforming Growth Factor beta2; D000205:Actomyosin; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020742:rhoA GTP-Binding Protein; C032881:lysophosphatidic acid