title: Tissue transglutaminase is an important player at the surface of human endothelial cells: evidence for its externalization and its colocalization with the beta(1) integrin.
abstract: We recently reported [J. Cell Sci. 110, 2461-2472 (1997)] that reduced expression of tissue transglutaminase (tTgase, type II) in human endothelial cell line ECV304 led to impaired cell spreading and adhesion; however, there is no immunocytochemical evidence for its presence and specific location at the surface of these cells. In this report we have stably transfected the same cell line with the cDNA for human tTgase which has been tagged at the C-terminus of the encoded protein with a 12-amino-acid peptide from protein kinase C epsilon. Using antibodies directed against this epitope tag peptide we show for the first time using immunogold electron microscopy and fluorescent immunocytochemistry the presence of cell surface-related tTgase. In cells undergoing attachment and cell spreading the enzyme appears to be concentrated at cell adhesion points which are rich in beta(1) integrin, suggesting that these areas may be the initial focal points for enzyme externalization. In more spread and confluent cells the enzyme appears more diffusely distributed along the basal membrane, with increased concentrations found at areas of cell-cell and cell-substratum contact. These findings strengthen the argument for the enzyme's role in cell-matrix interactions.
authors.affiliation: Department of Life Sciences, Nottingham Trent University, Nottingham, United Kingdom.
authors.forename: C A
authors.initials: CA
authors.lastname: Gaudry
authors.forename: E
authors.initials: E
authors.lastname: Verderio
authors.forename: R A
authors.initials: RA
authors.lastname: Jones
authors.forename: C
authors.initials: C
authors.lastname: Smith
authors.forename: M
authors.initials: M
authors.lastname: Griffin
mesh_terms: D001483:Base Sequence; D002448:Cell Adhesion / Q000502:physiology; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology* / Q000648:ultrastructure; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D004730:Endothelium, Vascular / Q000378:metabolism* / Q000648:ultrastructure; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D019012:Integrin beta1 / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D016253:Microscopy, Immunoelectron; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D014162:Transfection; D011503:Transglutaminases / Q000235:genetics / Q000378:metabolism*
chemical_list: D017931:DNA Primers; D018076:DNA, Complementary; D019012:Integrin beta1; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011503:Transglutaminases; D019204:GTP-Binding Proteins