title: Antibody-based analysis reveals "filamentous vs. non-filamentous" and "cytoplasmic vs. nuclear" crosstalk of cytoskeletal proteins.
abstract: To uncover the molecular composition and dynamics of the functional scaffold for the nucleus, three fractions of biochemically-stable nuclear protein complexes were extracted and used as immunogens to produce a variety of monoclonal antibodies. Many helix-based cytoskeletal proteins were identified as antigens, suggesting their dynamic contribution to nuclear architecture and function. Interestingly, sets of antibodies distinguished distinct subcellular localization of a single isoform of certain cytoskeletal proteins; distinct molecular forms of keratin and actinin were found in the nucleus. Their nuclear shuttling properties were verified by the apparent nuclear accumulations under inhibition of CRM1-dependent nuclear export. Nuclear keratins do not take an obvious filamentous structure, as was revealed by non-filamentous cytoplasmic keratin-specific monoclonal antibody. These results suggest the distinct roles of the helix-based cytoskeletal proteins in the nucleus.
authors.affiliation: Graduate School of Biostudies, Kyoto University, Kyoto 606-8501, Japan. Electronic address: kumeta@lif.kyoto-u.ac.jp.
authors.forename: Masahiro
authors.initials: M
authors.lastname: Kumeta
authors.forename: Yuya
authors.initials: Y
authors.lastname: Hirai
authors.forename: Shige H
authors.initials: SH
authors.lastname: Yoshimura
authors.forename: Tsuneyoshi
authors.initials: T
authors.lastname: Horigome
authors.forename: Kunio
authors.initials: K
authors.lastname: Takeyasu
mesh_terms: D000911:Antibodies, Monoclonal / Q000032:analysis* / Q000276:immunology; D002467:Cell Nucleus / Q000378:metabolism*; D005591:Chemical Fractionation; D003593:Cytoplasm / Q000378:metabolism*; D003598:Cytoskeletal Proteins / Q000032:analysis* / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans
chemical_list: D000911:Antibodies, Monoclonal; D003598:Cytoskeletal Proteins
keywords: Cytoskeleton; EGFP; Keratin; LMB; Monoclonal antibody; NES; NLS; NPC; Nuclear matrix; Nuclear scaffold; Nuclear transport; enhanced-GFP; leptomycin B; nuclear export signal; nuclear localization signal; nuclear pore complex