title: Molecular characterization and tissue distribution of ZO-2, a tight junction protein homologous to ZO-1 and the Drosophila discs-large tumor suppressor protein.
abstract: ZO-1 is a 210-225-kD peripheral membrane protein associated with cytoplasmic surfaces of the zonula occludens or tight junction. A 160-kD polypeptide, designated ZO-2, was found to coimmunoprecipitate with ZO-1 from MDCK cell extracts prepared under conditions which preserve protein associations (Gumbiner, B., T. Lowenkopf, and D. Apatira. 1991. Proc. Natl. Acad. Sci. USA. 88: 3460-3464). We have isolated ZO-2 from MDCK cell monolayers by bulk coimmunoprecipitation with ZO-1 followed by electroelution from preparative SDS-PAGE gel slices. Amino acid sequence information obtained from a ZO-2 tryptic fragment was used to isolate a partial cDNA clone from an MDCK library. The deduced amino acid sequence revealed that canine ZO-2 contains a region that is very similar to sequences in human and mouse ZO-1. This region includes both a 90-amino acid repeat domain of unknown function and guanylate kinase-like domains which are shared among members of the family of proteins that includes ZO-1, erythrocyte p55, the product of the lethal(1)discs-large-1 (dlg) gene of Drosophila, and a synapse-associated protein from rat brain, PSD-95/SAP90. The dlg gene product has been shown to act as a tumor suppressor in the imaginal disc of the Drosophila larva, although the functions of other family members have not yet been defined. A polyclonal antiserum was raised against a unique region of ZO-2 and found to exclusively label the cytoplasmic surfaces of tight junctions in MDCK plasma membrane preparations, indicating that ZO-2 is a tight junction-associated protein. Immunohistochemical staining of frozen sections of whole tissue demonstrated that ZO-2 localized to the region of the tight junction in a number of epithelia, including liver, intestine, kidney, testis, and arterial endothelium, suggesting that this protein is a ubiquitous component of the tight junction. Double-label immunofluorescence microscopy performed on cryosections of heart, a nonepithelial tissue, revealed the presence of ZO-1 but no ZO-2 staining at the fascia adherens, a specialized junction of cardiac myocytes which has previously been shown to contain ZO-1 (Itoh, M., S. Yonemura, A. Nagafuchi, S. Tsukita, and Sh. Tsukita. 1991. J. Cell Biol. 115:1449-1462). Thus it appears that ZO-2 is not a component of the fascia adherens, and that unlike ZO-1, this protein is restricted to the epithelial tight junction.
authors.affiliation: Program in Cell and Development Biology, Harvard Medical School, Boston, Massachusetts 02115.
authors.forename: L A
authors.initials: LA
authors.lastname: Jesaitis
authors.forename: D A
authors.initials: DA
authors.lastname: Goodenough
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D004285:Dogs; D029721:Drosophila Proteins; D004727:Endothelium / Q000737:chemistry; D004848:Epithelium / Q000737:chemistry; D005455:Fluorescent Antibody Technique; D016147:Genes, Tumor Suppressor; D007301:Insect Hormones / Q000737:chemistry*; D007365:Intercellular Junctions / Q000737:chemistry*; D008565:Membrane Proteins / Q000032:analysis* / Q000737:chemistry* / Q000302:isolation & purification; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009206:Myocardium / Q000737:chemistry; D010750:Phosphoproteins / Q000737:chemistry*; D025521:Tumor Suppressor Proteins; D062826:Zonula Occludens-1 Protein; D062827:Zonula Occludens-2 Protein
chemical_list: D029721:Drosophila Proteins; D007301:Insect Hormones; D008565:Membrane Proteins; D010750:Phosphoproteins; C569042:TJP1 protein, human; C569046:TJP2 protein, human; C569043:Tjp1 protein, mouse; C569044:Tjp1 protein, rat; C569047:Tjp2 protein, mouse; D025521:Tumor Suppressor Proteins; D062826:Zonula Occludens-1 Protein; D062827:Zonula Occludens-2 Protein; C070350:dlg1 protein, Drosophila