DNA-mediated transport of the intermediate filament protein vimentin into the nucleus of cultured cells

A number of characteristic properties of intermediate filament (IF) protein s, such as nucleic acid-binding activity, affinity for histones and structu ral relatedness to transcription factors and nuclear matrix proteins, in co njunction with the tight association of Ifs with the nucleus, suggest that these proteins might also fulfill nuclear functions in addition to their st ructure-organizing and -stabilizing activities in the cytoplasm. Yet, cytop lasmic IF proteins do not possess nuclear localization signals. In a search for carriers capable of transporting the IF protein vimentin into the nucl eus, complexes of FITC-vimentin with various DNAs were microinjected into t he cytoplasm of cultured cells and the intracellular distribution of the pr otein was followed by confocal laser scanning microscopy. The single-strand ed oligodeoxyribonucleotides oligo(dG)(25), oligo[d(GT)(12)G] and oligo[d(G (3)T(2)A)(4)G] proved to be excellent nuclear carriers for vimentin. Howeve r, in fibroblasts, fluorescence-labeled vimentin taken up by the nuclei rem ained undetectable with affinity-purified, polyclonal anti-vimentin antibod y, whereas it was readily identifiable in the nuclei of microinjected epith elial cells in this way. Moreover, when FITC-vimentin was preinjected into fibroblasts and allowed to assemble into the endogenous vimentin filament s ystem, it was still transferred into the nucleus by post-injected oligo(dG) (25), although to a lesser extent. Superhelical circular DNAs, like pBR322, SV40 and mitochondrial DNA, were also characterized by considerable capaci ties for nuclear vimentin transport; these transport potentials were totall y destroyed by relaxation or linearization of the DNA molecules. Neverthele ss, certain linear double-stranded DNA molecules with a high affinity for v imentin Ifs, such as repetitive telomere and centromere or mobile long inte rspersed repeat (LINE) DNA, could carry FITC-vimentin into the nucleus. Thi s was also true for a 375 bp extrachromosomal linear DNA fragment which occ urs in the cytoplasm of mouse tumor cells and which is capable of immortali zing human lymphocytes. On the basis of these results, it appears very like ly that cellular and viral products of reverse transcription as well as oth er extrachromosomal DNAs, which are circular, superhelical and apparently s huttling between the cytoplasm and the nucleus (eccDNA), are constantly Loa ded with vimentin in vimentin-positive cells. Since such DNAs are considere d as markers of genomic instability, it is conceivable that vimentin direct ly participates as an architectural, chromatin-modifying protein in recombi natorial processes set off by these DNAs in the nucleus.