Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Intermediate Filaments”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 901 records · Page 50Linked to original sources
PubMed1989

Specific disruption of intermediate filaments and the nuclear lamina by the 19-kDa product of the adenovirus E1B oncogene.

The 19-kDa protein encoded within the adenovirus E1B gene is essential for transformation by adenovirus and for proper regulation of viral early gene transcription. In order to investigate the biological function of the 19-kDa E1B protein, vectors were constructed to produce the 19-kDa protein in mammalian cells under the direction of heterologous promoters. Surprisingly, during transient expression, the E1B 19-kDa protein specifically associated with and disrupted the organization of intermediate filaments and the nuclear lamina, without disturbing the organization of other cytoskeletal networks. These results directly demonstrate an effect of a viral transforming protein on the cytoskeleton and suggest a role for intermediate filaments and the nuclear lamina in modulation of viral gene expression and the process of oncogenic transformation.

Adenovirus Early Proteins↗
PubMed1986

Binding of microtubule-associated protein 2 and tau to the intermediate filament reassembled from neurofilament 70-kDa subunit protein. Its regulation by calmodulin.

Two major brain microtubule-associated proteins (MAPs), MAP2 and tau, were found to bind to the intermediate filaments reassembled from neurofilament 70-kDa subunit protein (= 70-kDa filaments). The binding was saturable. The apparent dissociation constant (KD) for the binding of MAP2 to the 70-kDa filaments was estimated to be 4.8 X 10(-7) M, and the maximum binding reached 1 mol of MAP2/approximately 30 mol of 70-kDa protein. The apparent KD for the tau binding was 1.6 X 10(-6) M, and the maximum binding was 1 mol of tau/approximately 3 mol of 70-kDa protein. It was also found that MAP2 and tau did not compete with each other for binding to the 70-kDa filaments. Most interestingly, calmodulin, a ubiquitous Ca2+-binding protein in eukaryotic cells, was found to inhibit the binding of MAP2 and tau to the 70-kDa filaments. The inhibition by calmodulin was regulated by changes in Ca2+ concentration around 10(-6) M, and was canceled by trifluoperazine, a calmodulin inhibitor.

Animals↗
PubMed1997

Transient expression of epidermal filaggrin in cultured cells causes collapse of intermediate filament networks with alteration of cell shape and nuclear integrity.

Filaggrin is an intermediate filament-associated protein (IFAP) that aggregates epidermal keratin filaments in vitro and is thought to perform a similar function during terminal differentiation in vivo. To test this function in living cells, we transiently expressed constructs encoding human filaggrin in both simple epithelial cells (COS-7) and rat keratinocytes. Scanning laser confocal microscopy showed that filaggrin-positive cells had collapsed keratin and vimentin intermediate filament (IF) networks, and that filaggrin partially co-localized with the IF networks. Filaggrin was also detected diffusely in the cytoplasm and nucleus. In contrast, when profilaggrin-like constructs, containing five filaggrin domains separated by the linker sequences, were expressed in cultured cells, immunoreactive granules formed. This finding is reminiscent of the insoluble nature of native profilaggrin that accumulates in keratohyalin granules in vivo, suggesting that the linker peptides (present in profilaggrin but not filaggrin) are important for granule formation. Cells expressing filaggrin also displayed disruption of the nucleus and the nuclear envelope; they rounded up and lost attachment to the substratum, in contrast to control cells over-expressing beta-galactosidase. This functional test of filaggrin in living cells supports its role in the reorganization and packing of keratin IF in epidermal differentiation. Moreover, the observed effects on cell morphology and nuclear integrity suggest that filaggrin may contribute to the form of apoptosis associated with terminal differentiation in epidermis.

Animals↗
PubMed1984

Cell-specific domains of glial- and muscle-type intermediate filament proteins. Immunoaffinity chromatography and immunoblotting study of GFA protein and desmin.

In order to localize the cell specific domains of glial- and muscle-type intermediate filaments, the purified subunits (bovine GFA protein and chicken desmin) were fragmented and the digests passed through immunoaffinity columns or stained by the immunoblotting procedure to determine which fragments reacted with the monospecific polyvalent antisera. The following fragments were found immunoreactive according to these criteria: 30 K (GFA) and 33 K (desmin) N-bromosuccinimide fragments (tryptophan cleavage); 35 K (GFA) and 39 K (desmin) 2-nitro-5-thiocyanobenzoic acid fragments (cysteine cleavage); 18 K (GFA) and 9 K (desmin) cyanogen bromide fragments. Fragmentation of GFA protein was also accomplished using proteolytic digestion with chymotrypsin and trypsin. Two resistant core polypeptides, one about 37 K and stable in the chymotryptic digests and one about 21 K and stable in the tryptic digests bound specifically to the immunoaffinity columns. The 21 K tryptic fragment was found to contain the 18 K cyanogen bromide fragment. The fragmentation patterns support recently published structural domain models for intermediate filament proteins. The immunochemical findings indicate that the immunoreactive regions of GFA protein are located in the aminoterminal region of the middle domain of these models (coil I), while they appear to be situated in the aminoterminal headpiece of the protein in the case of desmin.

Amino Acid Sequence↗
PubMed1984

Expression of intermediate filaments in established human lung cancer cell lines. An indicator of differentiation and derivation.

A panel of established human pulmonary cancer cell lines, representing the major histopathologic groups according to the World Health Organization (WHO) classification (WHO 1, squamous cell carcinoma; WHO 2, small cell carcinoma; WHO 3, adenocarcinoma; WHO 4, large cell carcinoma) were examined for their expression of various types of intermediate filaments in order to determine their phenotypic differences and to attempt to disclose their histogenetic origin. The cells were investigated with antibodies specific for cytokeratin, vimentin, and neurofilament polypeptides with both immunofluorescence microscopy and immunoblotting techniques. Squamous cell carcinoma and adenocarcinomas expressed cytokeratin in accordance with the epithelial nature of these tumors but not neurofilament polypeptides. Small cell carcinomas, on the other hand, were positive for neurofilaments but negative for keratin. In contrast to small cell carcinoma, adenocarcinoma, and squamous cell carcinoma, one cell line derived from large cell carcinoma appeared to express both neurofilaments and keratin. All cell lines studied also contained variable amounts of vimentin, a phenotypic characteristic obtained by many cells under in vitro conditions. The results demonstrate, in accordance with our earlier observations in vivo, a distinctly divergent expression of intermediate filament proteins in different types of lung cancers. The persistence of this phenotypic heterogeneity in vitro consolidates the use of cell cultures as useful models to study the biologic behavior and interrelationships of lung cancers. Based on the present studies, and taking into account the occurrence of mixed forms of lung cancers, we present a hypothetical scheme of the histogenetic derivation of different types of lung cancers.

Adenocarcinoma↗
PubMed2004

Intermediate filaments are dynamic and motile elements of cellular architecture.

Recent evidence showing that intermediate filaments (IFs) are dynamic, motile elements of the cytoskeletal repertoire of vertebrate cells has overturned the long-standing view that they simply form static 'space filling' cytoplasmic networks. In fact, many types of IF are now known to engage in a remarkable array of movements that are closely associated with their assembly, disassembly and subcellular organization. Some of these motile properties are intrinsic to IFs and others are attributable to molecular crosstalk with either microtubules or actin-containing microfilaments. This crosstalk is, to a large extent, mediated by molecular motors, including conventional kinesin and cytoplasmic dynein. These motors are responsible for the high-speed delivery of nonfilamentous IF precursors and short filaments to specific regions of the cytoplasm, where they assemble into long IFs. Interestingly, the patterns and speeds of IF movements vary in different cell types and even within different regions of the same cell. These differences in motility may be related to their interactions with different types of molecular motor and/or other factors, such as IF-associated proteins.

Actin Cytoskeleton↗
PubMed2001

Intermediate filaments modulation in an in vitro model of the hepatic stellate cell activation or conversion into the lipocyte phenotype.

Hepatic stellate cells are intralobular connective tissue cells expressing the myofibroblast or the lipocyte phenotypes. They participate in homeostasis of the liver extracellular matrix, repair, regeneration, and fibrosis under the former phenotype, and control the retinol metabolism, storage, and release under the latter one. They are heterogeneous in terms of their tissue distribution, function, and expression of cytoskeletal proteins. We have studied the expressions of intermediate filaments in the cloned GRX cell line representative of murine hepatic stellate cells, by immunolabeling, reverse transcription polymerase chain reaction (RT-PCR), immunoprecipitation and Western blots. GRX cells expressed vimentin, desmin, glial fibrillary acidic protein (GFAP), and smooth muscle alpha actin (SM-alphaA). Vimentin, desmin, and SMN-alphaA were expressed in all cultures. GFAP showed a heterogeneous intensity of expression and did not form a filamentous cytoskeletal network, showing a distinct punctuate cytoplasmic distribution. When activated by inflammatory mediators, GRX cells increased expression of desmin and GFAP. Retinol-mediated induction of the lipocyte phenotype elicited a strong decrease of intermediate filament protein expression and the collapse of the filamentous structure of the cytoskeleton. Quiescent hepatic stellate precursors can respond to physiologic or pathologic stimuli, expressing activated myofibroblast or lipocyte phenotypes with distinct patterns of cytoskeleton structure, metabolic function, and interaction with the tissue environment.

Actins↗
PubMed2004

Intracellular assembly and sorting of intermediate filament proteins: role of the 42 amino acid lamin insert.

Nuclear and cytoplasmic intermediate filament (IF) proteins segregate into two independent cellular networks by mechanisms that are poorly understood. We examined the role of a 42 amino acid (aa) insert unique to vertebrate lamin rod domains in the coassembly of nuclear and cytoplasmic IF proteins by overexpressing chimeric IF proteins in human SW13+ and SW13- cells, which contain and lack endogenous cytoplasmic IF proteins, respectively. The chimeric IF proteins consisted of the rod domain of human nuclear lamin A/C protein fused to the amino and carboxyl-terminal domains of the mouse neurofilament light subunit (NF-L), which contained or lacked the 42 aa insert. Immunofluorescence microscopy was used to follow assembly and targeting of the proteins in cells. Chimeric proteins that lacked the 42 aa insert colocalized with vimentin, whereas those that contained the 42 aa insert did not. When overexpressed in SW13- cells, chimeric proteins containing the 42 aa formed very short or broken cytoplasmic filaments, whereas chimeric proteins that lacked the insert assembled efficiently into long, stable cytoplasmic filaments. To examine the roles of other structural motifs in intracellular targeting, we added two additional sequences to the chimera, a nuclear localization signal (NLS) and a CAAX motif, which are found in nuclear IF proteins. Addition of an NLS alone or an NLS in combination with the CAAX motif to the chimera with the 42 aa insert resulted in cagelike filament that assembled close to the nuclear envelope and nuclear lamina-like targeting, respectively. Our results suggest that the rod domains of eukaryotic nuclear and cytoplasmic IF proteins, which are related to each other, are still compatible upon deletion of the 42 aa insert of coassembly. In addition, NF-L end domains can substitute for the corresponding lamin domains in nuclear lamina targeting.

Amino Acid Sequence↗
PubMed1983

Intermediate filaments of the vimentin and prekeratin type in human epidermis.

Monospecific antibodies to intermediate filaments of mesenchymally derived vimentin, applied to frozen sections of human skin, specifically stained dendritic basal and suprabasal cells. Cellular morphology, distribution, reaction pattern with anti-HLA-DR in serial sections, and OKT6 in double-staining procedures identified these cells as melanocytes and Langerhans cells. As shown previously in animals keratinocytes stained with antiprekeratin exclusively. Thus, vimentin is an intracellular marker for Langerhans cells and melanocytes, differentiating them from keratinocytes.

Cytoskeleton↗
PubMed1988

Ubiquitin is a common factor in intermediate filament inclusion bodies of diverse type in man, including those of Parkinson's disease, Pick's disease, and Alzheimer's disease, as well as Rosenthal fibres in cerebellar astrocytomas, cytoplasmic bodies in muscle, and mallory bodies in alcoholic liver disease.

Polyclonal antibodies were raised which have a high affinity for conjugated ubiquitin. Immunocytochemistry was performed on paraffin sections of tissues showing well-characterized inclusion bodies. Ubiquitin was found as a component of the intermediate filament inclusion bodies characteristic of several major diseases including Lewy bodies of Parkinson's disease, Pick bodies of Pick's disease, Mallory bodies of alcoholic liver disease, cytoplasmic bodies of a specific myopathy, and Rosenthal fibres within astrocytes. Ubiquitin was also present in the three histological lesions characteristic of Alzheimer's disease. These observations suggest a fundamental role for ubiquitin in the formation of intermediate filament inclusion bodies in man, and have implications regarding the pathogenesis of these important diseases.

Alzheimer Disease↗
PubMed2000

Contribution of intermediate filaments to cell stiffness, stiffening, and growth.

It has been shown previously that intermediate filament (IF) gels in vitro exhibit stiffening at high-applied stress, and it was suggested that this stiffening property of IFs might be important for maintaining cell integrity at large deformations (Janmey PA, Evtenever V, Traub P, and Schliwa M, J Cell Biol 113: 155-160, 1991). In this study, the contribution of IFs to cell mechanical behavior was investigated by measuring cell stiffness in response to applied stress in adherent wild-type and vimentin-deficient fibroblasts using magnetic twisting cytometry. It was found that vimentin-deficient cells were less stiff and exhibited less stiffening than wild-type cells, except at the lowest applied stress (10 dyn/cm(2)) where the difference in the stiffness was not significant. Similar results were obtained from measurements on wild-type fibroblasts and endothelial cells after vimentin IFs were disrupted by acrylamide. If, however, cells were plated over an extended period of time (16 h), they exhibited a significantly greater stiffness before than after acrylamide, even at the lowest applied stress. A possible reason could be that the initially slack IFs became fully extended due to a high degree of cell spreading and thus contributed to the transmission of mechanical stress across the cell. Taken together, these findings were consistent with the notion that IFs play important roles in the mechanical properties of the cell during large deformation. The experimental data also showed that depleting or disrupting IFs reduced, but did not entirely abolish, cell stiffening. This residual stiffening might be attributed to the effect of geometrical realignment of cytoskeletal filaments in the direction of applied load. It was also found that vimentin-deficient cells exhibited a slower rate of proliferation and DNA synthesis than wild-type cells. This could be a direct consequence of the absence of the intracellular IFs that may be necessary for efficient mediation of mechanical signals within the cell. Taken together, results of this study suggest that IFs play important roles in the mechanical properties of cells and in cell growth.

Animals↗
PubMed2001

Desmuslin, an intermediate filament protein that interacts with alpha -dystrobrevin and desmin.

Dystrobrevin is a component of the dystrophin-associated protein complex and has been shown to interact directly with dystrophin, alpha1-syntrophin, and the sarcoglycan complex. The precise role of alpha-dystrobrevin in skeletal muscle has not yet been determined. To study alpha-dystrobrevin's function in skeletal muscle, we used the yeast two-hybrid approach to look for interacting proteins. Three overlapping clones were identified that encoded an intermediate filament protein we subsequently named desmuslin (DMN). Sequence analysis revealed that DMN has a short N-terminal domain, a conserved rod domain, and a long C-terminal domain, all common features of type 6 intermediate filament proteins. A positive interaction between DMN and alpha-dystrobrevin was confirmed with an in vitro coimmunoprecipitation assay. By Northern blot analysis, we find that DMN is expressed mainly in heart and skeletal muscle, although there is some expression in brain. Western blotting detected a 160-kDa protein in heart and skeletal muscle. Immunofluorescent microscopy localizes DMN in a stripe-like pattern in longitudinal sections and in a mosaic pattern in cross sections of skeletal muscle. Electron microscopic analysis shows DMN colocalized with desmin at the Z-lines. Subsequent coimmunoprecipitation experiments confirmed an interaction with desmin. Our findings suggest that DMN may serve as a direct linkage between the extracellular matrix and the Z-discs (through plectin) and may play an important role in maintaining muscle cell integrity.

Amino Acid Sequence↗
PubMed1987

Characterization of intermediate filaments in PC12 cells.

A 57 kDa protein is the major polypeptide in intermediate filament (IF)-enriched cytoskeletal preparations obtained from the neuronal cell line PC12 (rat pheochromocytoma). Under the conditions used to assemble IF in vitro from other cultured cell lines, 10 nm filaments are formed after 2 cycles of disassembly-assembly from PC12 IF-enriched cytoskeletal preparations; the 57 kDa protein is the major component of the final IF pellet. The 57 kDa protein is immunologically related to the BHK-21 fibroblast 55 kDa protein (vimentin), but a comparison of the peptide maps of PC12 57 kDa and BHK 55 kDa indicates that they are different proteins. With the use of a polyclonal antiserum to the PC12 57 kDa protein, immunofluorescence observations of PC12 cells not treated with NGF reveal a juxtanuclear "knot"-like structure. After NGF treatment, the "knots" are less prominent and many IF arrays are seen coursing through the cytoplasm and extending into the neurites. These immunofluorescence observations of the distribution of IF are corroborated by fine-structural analyses. SDS-PAGE analyses indicate that IF-enriched cytoskeletons isolated from NGF-treated cells have a polypeptide composition similar to that of untreated cells, that is, the 57 kDa protein remains the major polypeptide. SDS-PAGE and immunoblotting analyses show that untreated and NGF-treated PC12 cells also contain relatively minor amounts of the 68, 150, and 200 kDa neurofilament triplet (NFT) proteins. Under immunofluorescence, only 5% of untreated PC12 cells are found to contain a juxtanuclear "knot" labeled with NFT antibodies, but with time following NGF treatment, the number of fluorescent cells increases. After about 2 weeks of NGF treatment, all of the PC12 cells appear to contain NFT antibody-positive filamentous structures. As assessed by immunofluorescence, the NFT polypeptides appear to codistribute with the 57 kDa protein in both untreated and NGF-treated PC12 cells. These data indicate that PC12 cells contain IF composed of a complex set of polypeptides, including a previously unidentified 57 kDa IF protein. While NGF may induce production of NFT polypeptides, there does not appear to be a "switch" from known mesenchymal IF polypeptide expression to NFT polypeptide expression upon stimulation of PC12 cells with NGF.

Adrenal Gland Neoplasms↗
PubMed1995

Intermediate filament expression and lectin histochemical features of canine transmissible venereal tumour.

Immunocharacterization of intermediate filament proteins and lectin-binding studies were carried out in canine transmissible venereal tumour (TVT), a unique neoplasia sharing some epidemiological features with Kaposi's sarcoma in humans. Neoplastic cells express vimentin, but neither cytokeratin nor desmin. Regarding lectins, TVT cells express receptors for Triticum vulgaris (WGA), Concanavalia ensiformis (Con A) and Ricinus communis I (RCA-I). They appear to be negative for Ulex europaeus-I (UEA-I), Arachis hypogaea (PNA), Glycine maximus (SBA) and Dolichos biflorus (DBA).

Animals↗
PubMed2003

Functional complexity of intermediate filament cytoskeletons: from structure to assembly to gene ablation.

The cell biology of intermediate filament (IF) proteins and their filaments is complicated by the fact that the members of the gene family, which in humans amount to at least 65, are differentially expressed in very complex patterns during embryonic development. Thus, different tissues and cells express entirely different sets and amounts of IF proteins, the only exception being the nuclear B-type lamins, which are found in every cell. Moreover, in the course of evolution the individual members of this family have, within one species, diverged so much from each other with regard to sequence and thus molecular properties that it is hard to envision a unifying kind of function for them. The known epidermolytic diseases, caused by single point mutations in keratins, have been used as an argument for a role of IFs in mechanical "stress resistance," something one would not have easily ascribed to the beaded chain filaments, a special type of IF in the eye lens, or to nuclear lamins. Therefore, the power of plastic dish cell biology may be limited in revealing functional clues for these structural elements, and it may therefore be of interest to go to the extreme ends of the life sciences, i.e., from the molecular properties of individual molecules including their structure at the atomic level to targeted inactivation of their genes in living animals, mouse, and worm to define their role more precisely in metazoan cell physiology.

Animals↗
PubMed1991

Quantitative and qualitative alterations of neuronal and glial intermediate filaments in rat nervous system after exposure to 2,5-hexanedione.

The precise mechanism for the neurotoxicity of 2,5-hexanedione is not known, but cross-linking of neurofilament proteins has been suggested as one possibility. In this study the effects of long-term exposure to 2,5-hexanedione were studied in the rat nervous system with special reference to regional changes in the quantities of neuronal and glial intermediate filaments. Using enzyme-linked immunosorbent assays the concentrations of 68- and 200-kDa neurofilament polypeptides were shown to be reduced in all brain regions studied. Similar results were obtained in the sciatic nerve. The concentration of glial fibrillary acidic protein was decreased in the cerebellar vermis and the dorsal cerebral cortex, whereas it was increased in the spinal cord, a result suggesting a regional variation in glial sensitivity. The intermediate filaments of the exposed animals were also immunoblotted using polyclonal antisera against the various neurofilament polypeptides and glial fibrillary acidic protein. In all tissues studied, several aggregates with molecular weights higher than those of the monomeric polypeptides were demonstrated. Contrary to clinical observations, these data indicate pronounced effects in both CNS and PNS and call for further studies on CNS effects in humans.

Animals↗
PubMed1982

The cDNA sequence of a human epidermal keratin: divergence of sequence but conservation of structure among intermediate filament proteins.

We have determined the DNA sequence of a cloned cDNA that is complementary to the mRNA for the 50 kilodalton (kd) human epidermal keratin. This provides the first amino acid sequence for a cytoskeletal keratin. Comparison of this sequence with those of other keratins reveals an evolutionary relationship between the cytoskeletal and the microfibrillar keratins, but shows no homology to matrix or feather keratins. The 50 kd keratin shares 28%-30% homology with partial sequences of other intermediate filament proteins, which suggests that keratins may be the most distantly related members of this class of fibrous proteins. Our computer analyses predict that the 50 kd keratin contains two long alpha-helical domains separated by a cluster of helix-inhibitory residues in the middle of the protein. These findings indicate that despite major sequence divergence among intermediate filament proteins, they retain sequences compatible with secondary structural features that appear to be common to all of them.

Amino Acid Sequence↗
PubMed1994

Intermediate filament and related proteins: potential activators of nucleosomes during transcription initiation and elongation?

Intermediate filament (IF) protein tetramers contain two DNA- and core-histone-binding motifs in rotational symmetry in one and the same structural entity. We propose that IF protein oligomers might displace histone octamers from nucleosomes in the process of transcription initiation and elongation, to deposit them transiently on their alpha-helical coiled-coil domains. We further propose that structurally related proteins of the karyoskeleton, constructed from an alpha-helical domain capable of coiled-coil formation and a basic DNA-binding region adjacent to it, may be similarly involved in nucleosome activation. These proteins would function as auxiliary factors that disrupt nucleosomal structure to permit transcription and other DNA-dependent processes to proceed expiditiously.

Animals↗