Interaction in vitro of Non-Epithelial Intermediate Filament Proteins with Histones

Abstract

Non-epithelial intermediate filament (IF) subunit proteins show a high and specific affinity for core histones at physiological ionic strength. When IF proteins are titrated with a mixture of core histones and linker histone H1, in general the latter is totally excluded from com plexation and in the adducts formed the moderately-arginine-rich histones H2A and H2B are progressively replaced by the very-arginine-rich histones H3 and H4. At histone saturation, 2 molecules of nonneuronal IF protein bind 1 histone HI molecule or 8 core histone m olecules, whereas due to its glutamic acid rich, C terminal extensions one dimer of thp 68 kD npnrofilament nrotein associates with 3 molecules of histone H1 or 24 molecules of core histones. The salt stability of the insoluble association products is dependent on the amount and arginine content of the constituent histone species. Rem oval of the non-α-helical N- and C-terminal polypeptides from IF proteins by partial chymotryptic digestion does not affect their histone-binding characteristics. Since core histones are only partially inactivated by limited tryptic digestion, they also appear to react through their a-helix-rich central domains; the limit peptide derived from histone H1 is com pletely inactive at physiological ionic strength. Affinity chromatography of rod domains of IF proteins on core histone-Sepharose 4B and of histones and their limit peptides on vim entin-Sepharose 4B has shown that the interactions involving fractions of histones H3 and H4 are extrem ely resistant to salt and can be dissociated only with arginine or salt under denaturing conditions. In general, the experim ental results revealed close parallels between the association of histones with IF proteins and their interaction with DNA.

Isolation of SDS-Stable Complexes of the Intermediate Filament Protein Vimentin with Repetitive, Mobile, Nuclear Matrix Attachment Region, and Mitochondrial DNA Sequence Elements from Cultured Mouse and Human Fibroblasts

“…As pointed out previously (Tolstonog et al, 2000), IF proteins are comparable to members of the RAD52 epistasis group and replication protein A, which play an essential role in homologous recombination repair. Conversely, putting its affinity for single-stranded (Traub et al, 1983(Traub et al, , 1992, supercoiled (Kühn et al, 1987), triple-helical, and Z (unpublished observations) DNA as well as for core histones (Traub et al, 1986) into effect, vimentin will certainly also contribute to the marked vulnerability of nuclear matrix-associated DNA to damage by a multitude of noxious physical and chemical agents such as ionizing radiation, UV light, carcinogens, and ROS (reviewed in Boulikas, 1995). Via opening of chromatin structure, vimentin would thus exert opposing effects on the genome: one rendering it more accessible to mutagens and one allowing the invasion of constituents of the recombination/repair machinery at sites of DNA lesions.…”

“…On the basis of these observations and the strong affinity of vimentin for repetitive and mobile DNA sequences, especially if these elements are, in addition, under superhelical tension (Kühn et al, 1987), it has been proposed that vimentin also plays a role in recombination processes (Tolstonog et al, 2000), which take place, for instance, during cell immortalization or cell senescence (Macieira-Coelho, 1996; see also accompanying paper in this issue). As a DNA-binding protein with an additional strong and selective reactivity with core histones (Traub et al, 1986), vimentin may mediate these processes by acting as a chromatin-modifying scaffold protein.…”

InteractionIn Vitroof Type III Intermediate Filament Proteins with Z-DNA and B-Z-DNA Junctions

“…In this case, the cIF proteins may make use of their high affinities in vitro for such nuclear constituents as the lamin B component of the nuclear lamina Djabali et al, 1991;Bastos et al, 1992), DNAs and RNAs Vorgias and Traub, 1986;Tolstonog et al, 2000;Q. Wang et al, 2001), and histones (Traub et al, 1987b). That the affinity binding of type III IF proteins to a large variety of repetitive and mobile, recombinogenic DNA sequences (X.…”

“…Among the crossconnected DNA fragments, centromeric and telomeric DNA were the most prominent, in addition to micro-and minisatellites and transposable sequence elements of the LINE, SINE and A-type particle families. The specificity of the cIF protein-histone interactions emerged from the exclusion of linker histone H1 from complex formation as long as core histones were present in small excess, the preferential affinity of the filament proteins for the arginine-rich histone species H3 and H4, and the stoichiometry of the adducts, with 16 core histone molecules bound to each cIF protein tetramer (Traub et al, 1987b).…”

Amino-terminal Polypeptides of Vimentin Are Responsible for the Changes in Nuclear Architecture Associated with Human Immunodeficiency Virus Type 1 Protease Activity in Tissue Culture Cells

“…Critically important to the interpretation and conclusions of this study are the data obtained from control experiments: the organization of the nuclei of cells lacking vimentin remains unaffected by treatment with HIV-1 PR. Because the limit digest of vimentin by HIV-1 PR produces a mixture of several peptides (Shoeman et al, 1990a,b), corresponding roughly to the head domain, the rod domain and the tail domain, and because it has been shown that the various domains of vimentin can interact with themselves and some other related proteins (Traub et al, 1992b;Meng et al, 1996) as well as with histones (Traub et al, 1986), it was of interest to define which peptide(s) of this mixture was responsible for the changes observed. The amino-terminal peptide NT1 was originally chosen because it was available in sufficient quantity and purity, is almost identical to the largest amino-terminal peptide produced by HIV-1 PR, and because it encompasses a domain of vimentin known to be involved in interactions with a variety of compounds, including nucleic acids, lipids, and coiled-coil protein structures (Traub et al, 1992b and references therein).…”