title: Functional analysis of nuclear pore complex protein Nup62/p62 using monoclonal antibodies.
abstract: The nuclear pore complex (NPC) is an enormous structure embedded in the double membrane of the nuclear envelope that acts as a passageway for nucleocytoplasmic transport. The vertebrate NPC is comprised of about 30 unique proteins. Nup62/p62, a major component of the NPC, has been reported to interact directly with several nuclear transport factors, including importin-beta and NTF2. However, it has not been shown how the interaction of Nup62/p62 with transport factors is involved in nucleocytoplasmic transport. The present study reports on the preparation of monoclonal antibodies (MAbs) directed against human Nup62/p62 and a functional analysis of Nup62/p62 using antibodies in living cells. Hybridomas producing the antibodies were produced by the hybridization of mouse myeloma cells with medial iliac lymph node cells from an immunized rat. These MAbs specifically recognized Nup62/p62 as evidenced by immunoblotting analysis using a nuclear membrane fraction. In the immunostaining using MAbs, a punctuate nuclear rim staining pattern was observed. Moreover, cytoplasmic injected-anti-Nup62/p62 MAbs were rapidly targeted to the nuclear pore of cultured cells and some of them inhibited normal cell division, causing the formation of abnormal nuclei. The antibodies described in this study provide the means for immunochemical analyses of the NPC protein Nup62/p62 in mammalian cells, and represent useful molecular tools that should permit a better understanding of the biological roles and cellular dynamics of this protein in nucleocytoplasmic transport, cell division, and nuclear organization.
authors.affiliation: Department of Bioengineering, Graduate School of Engineering, Osaka City University, Sugimoto, Osaka, Japan.
authors.forename: Takaomi
authors.initials: T
authors.lastname: Fukuhara
authors.forename: Naoko
authors.initials: N
authors.lastname: Sakaguchi
authors.forename: Jun
authors.initials: J
authors.lastname: Katahira
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Yoneda
authors.forename: Kenji
authors.initials: K
authors.lastname: Ogino
authors.forename: Taro
authors.initials: T
authors.lastname: Tachibana
mesh_terms: D021581:Active Transport, Cell Nucleus; D000818:Animals; D000911:Antibodies, Monoclonal / Q000302:isolation & purification; D002455:Cell Division; D002467:Cell Nucleus / Q000502:physiology; D003593:Cytoplasm / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D008562:Membrane Glycoproteins / Q000276:immunology* / Q000502:physiology; D051379:Mice; D028861:Nuclear Pore Complex Proteins / Q000276:immunology* / Q000502:physiology; D051381:Rats
chemical_list: D000911:Antibodies, Monoclonal; D008562:Membrane Glycoproteins; D028861:Nuclear Pore Complex Proteins; C049312:nuclear pore protein p62