Properties of the nonhelical end domains of vimentin suggest a role in maintaining intermediate filament network structure.
To investigate the functional role of the nonhelical domains of the intermediate filament (IF) protein vimentin, we carried out transient transfection of constructs encoding fusion proteins of these domains with enhanced green fluorescent protein (EGFP). Expression of these fusion proteins did not have any effect on the endogenous IF networks of transfected cells. However, the head domain-EGFP fusion protein localized almost exclusively to the nucleus. This localization could be disrupted in a reversible fashion by chilling cells. Furthermore, the head domain was capable of targeting to the nucleus a strictly cytoplasmic protein, pyruvate kinase. Thus, the vimentin head domain contains information that specifically directs proteins into the nucleus. In contrast, the nonhelical tail domain of vimentin, when expressed as a fusion protein with EGFP, was retained in the cytoplasm. Cytoplasmic retention of tail domain-containing fusion proteins appeared to be dependent on the integrity of the microtubule network. Our results are consistent with a proposal that the nonhelical end domains of vimentin are involved in maintaining an extended IF network by exerting oppositely directed forces along the filaments. The head domains exert a nuclear-directed force while the tail domains extend the IF network toward the cell periphery via a microtubule-dependent mechanism.
Lowrie
D J
DJ
Department of Cell Biology, Neurobiology, and Anatomy, University of Cincinnati College of Medicine, Cincinnati, Ohio 45267-0521, USA.
Stickney
J T
JT
Ip
W
W
D021581:Active Transport, Cell Nucleus; D000818:Animals; D002467:Cell Nucleus; D003593:Cytoplasm; D017930:Genes, Reporter; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D006801:Humans; D007382:Intermediate Filaments / Q000737:chemistry* / Q000648:ultrastructure; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008856:Microscopy, Fluorescence; D017434:Protein Structure, Tertiary; D011770:Pyruvate Kinase / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured; D014746:Vimentin / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology
D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D014746:Vimentin; D049452:Green Fluorescent Proteins; D011770:Pyruvate Kinase