Biomedical subjects

G Wiche

Publications and source records attributed to G Wiche.

At least 73 records · Page 4Linked to original sources

PubMed1989

An antigenic profile of Lewy bodies: immunocytochemical indication for protein phosphorylation and ubiquitination.

An antigenic profile of subcortical and cortical Lewy bodies was determined in the presence or absence of neurofibrillary tangles in the same brain using antisera and monoclonal antibodies to various cytoskeletal elements as well as to determinants not present in the normal cytoskeleton. The cores of many Lewy bodies were strongly reactive with a monoclonal antibody to paired helical filaments which has been shown to recognize ubiquitin. This antibody also stained Marinesco bodies in the same tissue sections. Two monoclonal antibodies to phosphorylated epitopes of neurofilament proteins (SM I 31, SM I 34) stained the peripheries of about 40% of all discernable Lewy bodies on untreated paraffin sections. Reactivity with a monoclonal antibody to neurofilaments (SM I 33) appeared only after pretreatment of the sections with phosphatase. Lewy bodies did not bind antibodies to tau protein. Our results show that, as previously shown for neurofibrillary tangles, Lewy bodies also contain ubiquitin. The uncovering of neurofilament epitopes by treatment with phosphatase indicates that abnormal phosphorylation of cytoskeletal elements may play a role in the pathogenesis of the Lewy body.

Aged↗

PubMed1989

Plectin: general overview and appraisal of its potential role as a subunit protein of the cytomatrix.

Plectin has recently been identified as a widespread and abundant cytoplasmic protein of mammalian cells. In this article the available data on plectin are reviewed, focusing on plectin's occurrence and localization in various cell types and tissues, its biochemical characterization, and its molecular interaction partners. Furthermore, the putative role of this protein has a multifunctional connecting link of the cytomatrix and its structural as well as functional relationship to other cytoskeletal proteins is discussed. It is concluded that plectin is potentially the most versatile crosslinking element of the cytomatrix reported to date.

Animals↗

PubMed1988

Cytoskeleton-associated plectin: in situ localization, in vitro reconstitution, and binding to immobilized intermediate filament proteins.

The association and interaction of plectin (Mr 300,000) with intermediate filaments and filament subunit proteins were studied. Immunoelectron microscopy of whole mount cytoskeletons from various cultured cell lines (rat glioma C6, mouse BALB/c 3T3, and Chinese hamster ovary) and quick-frozen, deep-etched replicas of Triton X-100-extracted rat embryo fibroblast cells revealed that plectin was primarily located at junction sites and branching points of intermediate filaments. These results were corroborated by in vitro recombination studies using vimentin and plectin purified from C6 cells. Filaments assembled from mixtures of both proteins were extensively crosslinked by oligomeric plectin structures, as demonstrated by electron microscopy of negatively stained and rotary-shadowed specimens as well as by immunoelectron microscopy; the binding of plectin structures on the surface of filaments and cross-link formation occurred without apparent periodicity. Plectin's cross-linking of reconstituted filaments was also shown by ultracentrifugation experiments. As revealed by the rotary-shadowing technique, filament-bound plectin structures were oligomeric and predominantly consisted of a central globular core region of 30-50 nm with extending filaments or filamentous loops. Solid-phase binding to proteolytically degraded vimentin fragments suggested that plectin interacts with the helical rod domain of vimentin, a highly conserved structural element of all intermediate filament proteins. Accordingly, plectin was found to bind to the glial fibrillar acidic protein, the three neurofilament polypeptides, and skin keratins. These results suggest that plectin is a cross-linker of vimentin filaments and possibly also of other intermediate filament types.

Animals↗

PubMed1987

Structure and hydrodynamic properties of plectin molecules.

Plectin is a cytoskeletal, high molecular weight protein of widespread and abundant occurrence in cultured cells and tissues. To study its molecular structure, the protein was purified from rat glioma C6 cells and subjected to chemical and biophysical analyses. Plectin's polypeptide chains have an apparent molecular weight of 300,000, as shown by one-dimensional sodium dodecyl sulfate/polyacrylamide electrophoresis. Cross-linking of non-denatured plectin in solution with dimethyl suberimidate and electrophoretic analyses on sodium dodecyl sulfate/agarose gels revealed that the predominant soluble plectin species was a molecule of 1200 X 10(3) Mr consisting of four 300 X 10(3) Mr polypeptide chains. Hydrodynamic properties of plectin in solution were obtained by sedimentation velocity centrifugation and high-pressure liquid chromatography analysis yielding a sedimentation coefficient of 10 S and a Stokes radius of 27 nm. The high f/fmin ratio of 4.0 indicated a very elongated shape of plectin molecules and an axial ratio of about 50. Shadowing and negative staining electron microscopy of plectin molecules revealed multiple domains: a rigid rod of 184 nm in length and 2 nm in diameter, and two globular heads of 9 nm diameter at each end of the rod. Circular dichroism spectra suggested a composition of 30% alpha-helix, 9% beta-structure and 61% random coil or aperiodic structure. The rod-like shape, the alpha-helix content as well as the thermal transition within a midpoint of 45 degrees C and the transition enthalpy (168 kJ/mol) of secondary structure suggested a double-stranded, alpha-helical coiled coil rod domain. Based on the available data, we favor a model of native plectin as a dumb-bell-like association of four 300 X 10(3) Mr polypeptide chains. Electron microscopy and turbidity measurements showed that plectin molecules self-associate into various oligomeric states in solutions of nearly physiological ionic strength. These interactions apparently involved the globular end domains of the molecule. Given its rigidity and elongated shape, and its tendency towards self-association, plectin may well be an interlinking element of the cytoskeleton that may also form a network of its own.

Animals↗

PubMed1987

Plectin from bovine lenses. Chemical properties, structural analysis and initial identification of interaction partners.

Plectin was purified to near homogeneity from epithelial and cortical cell layers of bovine lenses using a simple and fast purification scheme that included as last step, gel permeation chromatography in the presence of 0.25% sodium N-lauroyl sarcosinate. Lens cell plectin showed extensive structural homology to plectin from cultured cells as revealed by immunoblotting experiments and amino acid analysis. Further characterization included solubility in various buffer solutions, codistribution with vimentin in repeated rounds of intermediate filament disassembly and assembly, and hydrodynamic behaviour in high-performance gel permeation chromatography. Electron microscopy of negatively stained and rotary shadowed plectin molecules revealed a dumb-bell-like structure with an estimated relative molecular mass of 1.16 X 10(6). Specific head-to-head self-interaction of plectin molecules at low salt concentrations and formation of large aggregates under high-salt and physiological conditions was also demonstrated. Isolation, as well as reconstitution of soluble protein complexes containing plectin, vimentin and other cytoskeletal and membrane skeleton proteins, provided first hints to plectin's role as an interlinking component of the cytoskeleton and the membrane skeleton of lens tissue.

Amino Acids↗

PubMed1987

Plectin and IFAP-300K are homologous proteins binding to microtubule-associated proteins 1 and 2 and to the 240-kilodalton subunit of spectrin.

Structural and functional characteristics of plectin from intermediate filament preparations of rat glioma C6 cells were compared to those of the intermediate filament-associated protein of Mr = 300,000 (IFAP-300K) of baby hamster kidney cells (Yang, H.-S., Lieska, N., Goldman, A.E., and Goldman, R.D. (1985) J. Cell Biol. 100, 620-631). After radiolabeling and proteolytic digestion under varied conditions, both proteins yielded nearly identical peptide maps. Immunological cross-reactivity, co-migration on one- and two-dimensional high-resolution gels, chromatofocusing, and amino acid analysis demonstrated structural homology as well. In vivo labeling with 32Pi showed that plectin was the target for cAMP-independent protein kinases which phosphorylated 18-kDa domains at the end(s) of the molecule. Previously reported phosphorylation sites for cAMP-dependent and a newly identified site for Ca2+/calmodulin-dependent protein kinases were located on different domains. In solid-phase binding assays, plectin bound to vimentin, microtubule-associated proteins 1 and 2, the 240-kDa chain of brain fodrin, and alpha-spectrin from human erythrocytes. Similar characteristics were revealed for corresponding 300-kDa components of various other cell lines, supporting the concept that plectin is a general cytoskeletal cross-linking element, probably of multiple function.

Animals↗

PubMed1987

Neurofibrillary tangles in Alzheimer's disease and progressive supranuclear palsy: antigenic similarities and differences. Microtubule-associated protein tau antigenicity is prominent in all types of tangles.

The antigenic profile of neurofibrillary tangles (NFT) in Alzheimer's disease (AD), senile dementia of Alzheimer type (SDAT), progressive supranuclear palsy (PSP) and in non-demented aged humans was investigated by light and electron microscopic immunocytochemistry using antisera and monoclonal antibodies to tubulin, microtubule-associated proteins (MAP1, MAP2 and tau), neurofilament proteins and determinants unique to Alzheimer paired helical filaments (PHF). Antibodies to tau proteins labeled NFT in all cases investigated (AD, SDAT, PSP and non-demented aged humans). However, one monoclonal antibody to PHF recognized numerous tangles in AD/SDAT, but only a small minority of the PSP tangles. Antibodies to tubulin, MAP1, MAP2 and neurofilament proteins did not selectively stain NFT. Whereas pretreatment of sections with phosphatase was required for the detection of tangles with Tau-1 monoclonal antibody, digestion of sections with either phosphatase or pronase had no significant effect on the staining pattern obtained with the other antibodies. Our studies show that, as previously described for AD/SDAT, phosphorylated tau polypeptides are also a major antigenic determinant of tangles in PSP, indicating that tangle formation may follow a common pathogenetic pathway in neurofibrillary degenerations. There is, however, at least one epitope in AD/SDAT tangles which seems to be absent on, or at least inaccessible in, the 15-nm straight fibrils of PSP.

Adult↗

PubMed1987

Binding specificities of purified porcine brain alpha- and beta-tubulin subunits and of microtubule-associated proteins 1 and 2 examined by electron microscopy and solid-phase binding assays.

In this study, the molecular interaction of separated alpha- and beta-tubulin with purified microtubule-associated protein 1 (MAP 1) and MAP 2 was studied using electron microscopy and solid-phase binding assays with 125I-radiolabeled proteins. Electron microscopy of proteins recovered from sodium dodecyl sulfate polyacrylamide gels and subsequently incubated in various combinations under conditions promoting tubulin polymer formation revealed that both subunits have binding sites for MAP 1 as well as MAP 2. Overlays of nitrocellulose-transblotted MAPs with electrophoretically separated tubulin subunits eluted from gels confirmed these results. In overlays of nitrocellulose-immobilized tubulin subunits with gel-eluted MAP 2, self-association of MAP 2, but no binding to tubulin was detected. However, overlays with MAP 1 and MAP 2 purified under nondenaturing conditions revealed binding of both MAPs to beta-tubulin. In addition, these experiments demonstrated binding of both MAPs to MAP 2 and to the neurofilament proteins NF 70, NF 150 and NF 200. It is concluded that both alpha- and beta-tubulin possess binding sites for MAP 1 as well as MAP 2, but that the accessibility and/or binding affinity of these sites are strongly dependent on the tertiary structure of proteins. The demonstrated in vitro binding of MAP 1 and MAP 2 to all three neurofilament proteins as well as to MAP 2 confirms their presumed role as cytoskeletal linking proteins.

Animals↗

PubMed1986

Molecular aspects of MAP-1 and MAP-2: microheterogeneity, in vitro localization and distribution in neuronal and nonneuronal cells.

We have studied various aspects of MAP-1 and MAP-2 from neuronal as well as nonneuronal sources. MAP-1 and MAP-2 polymerized from brain were resolved into a number of subcomponents upon electrophoresis on low percentage gels. Based on peptide mappings performed under a variety of different conditions, we conclude that the three major subcomponents of MAP-1 have very similar, though not identical structures. The two major MAP-2 subcomponents might have identical structure, because their peptide maps were hardly distinguishable. The apparent microheterogeneity of high Mr MAPs is not yet understood on a molecular basis. Proteolysis during isolation or a different degree of phosphorylation, however, seems to be an unlikely cause for microheterogeneity. When localized on microtubules polymerized in vitro by electron microscopy, both MAP-1 and MAP-2 polypeptides apparently form helical arrays on the polymer's surface with periodicities of 100 nm. In the presence of taxol, MAPs form irregular and bulky extensions. Both MAPs are found to be widespread in neuronal as well as nonneuronal cells. MAP-1- and MAP-2-related polypeptides, together with other high Mr proteins, such as plectin, were associated with microtubules polymerized by taxol from extracts of a nonneuronal cultured cell line. MAP-2 from cultured cells was found to be extremely sensitive to proteolysis, in particular in the presence of free Ca-ions. MAP-1 and MAP-2 generally were found associated with typical microtubule structures such as interphase and spindle microtubules and primary cilia. A differential distribution of MAP-1 and MAP-2 was clearly evident in neural tissues, where MAP-2 was restricted to cell bodies and dendrites, whereas MAP-1 was present also in axons. Moreover, a differential distribution of MAPs and tubulin was observed in de-and regenerating peripheral nerve, and in a few occasions, also with nonneuronal cells. A quite unexpected result was the identification of a protein in the extracellular matrix of cultured fibroblast cells, which has antigenic determinants in common with MAP-1 and MAP-2 from brain. As a whole, the data presented support a concept in which a family of structurally homologous, though not identical, high Mr polypeptides constitute the crosslinking elements between microtubules and various other cellular components. The structural diversity of these polypeptides might play a role in the development and dynamic changes in the cytoskeletal architecture.

Animals↗

PubMed1985

Microheterogeneity of microtubule-associated proteins, MAP-1 and MAP-2, and differential phosphorylation of individual subcomponents.

High molecular weight microtubule-associated proteins 1 and 2 (MAP-1 and MAP-2), prepared by copolymerization with tubulin, were electrophorectically separated into three and two major subcomponents, respectively, using 5% sodium dodecyl sulfate-polyacrylamide gels. By two-dimensional gel electrophoresis, all five MAP components were shown to possess a pI of around 5. Four of these proteins, MAP-1A, MAP-1C, MAP-2A, and MAP-2B, present in comparable amounts, were iodinated after electrophoretic separation and analyzed by two-dimensional peptide mapping. With both trypsin and V8 protease, almost identical patterns were obtained from MAP-2A and MAP-2B. MAP-1A and MAP-1C, too, gave similar digestion patterns, although some differences were noted. Incubation with [gamma-32P]ATP demonstrated that endogeneous protein kinase activities phosphorylated individual subcomponents at different rates. MAP-2A, the highest labeled component, was phosphorylated 2.5-fold compared to MAP-2B both in the presence and the absence of cAMP. Labeling of MAP-1 subcomponents was 4 times less than that of MAP-2A in the absence and 16 times less in the presence of cAMP. 32P-labeled MAP-2A and MAP-2B bands were indistinguishable by one-dimensional peptide mapping, as were the three MAP-1 bands. For both MAP-1 and MAP-2 subcomponents, cAMP induced phosphorylation at new molecular sites. Incubation of radiolabeled microtubule proteins with 1 mM ATP effected, upon electrophoresis, a clear shift of MAP-2A and MAP-2B bands to positions of higher apparent molecular weights, while only slightly affecting MAP-1 bands.

Adenosine Triphosphate↗

PubMed1985

Mr 205,000 sulfoglycoprotein in extracellular matrix of mouse fibroblast cells is immunologically related to high molecular weight microtubule-associated proteins.

Rabbit antibodies raised against microtubule-associated protein 1 (MAP-1) from hog brain were found to crossreact with extracellular matrix components of mouse BALB/c 3T3 cell cultures. As shown by immunofluorescence microscopy of confluent cell cultures, the extracellular MAP-related antigen was located on dense fibrillar network arrays underlying and surrounding the cells. The immunoreactive material was sensitive to trypsin but resistant to collagenase. The microtubule-disrupting drug colcemid had no visible effect on the morphology of the anti-MAP-stained network, whereas treatment with cytochalasin B provoked its abolishment. Simian virus 40-transformed BALB/c 3T3 cells expressed considerably less extracellular antigen than did the nontransformed cells. After in vivo radiolabeling of BALB/c 3T3 cells, a secreted polypeptide of Mr 205,000 was isolated by immuno-precipitation from culture media as well as from cell-free extracellular matrices. This antigen was identified as a sulfoglycoprotein, noncollageneous in nature, that undergoes intermolecular disulfide bonding. Anti-MAP-1 antibodies affinity-purified on the extracellular Mr 205,000 protein were immunoreactive with MAP-1 and MAP-2 from brain and decorated cytoplasmic microtubules as demonstrated by immunoblotting and immunofluorescence microscopy. Thus, a structural relationship between cytoskeletal and extracellular polypeptides is demonstrated.

Animals↗

PubMed1985

Monoclonal antibody to a 43 000 Mr surface protein of a human leukaemia cell line (THP-1) crossreacts with the fibroblast intermediate filament protein vimentin.

Monoclonal antibodies were produced against surface antigens of live cells from a human acute monocytic leukaemia cell line (THP-1). One clone, VIC-C2, when assayed by immunofluorescence microscopy, brightly stained the surface of THP-1 cells and the cytoplasm of Langerhans cells, fibroblasts and melanocytes in sections of human skin. The immunoreactive cytoplasmic structures were filamentous and resembled intermediate filaments. By double immunofluorescence microscopy using VIC-C2 and polyclonal antibodies to vimentin, the VIC-C2 antigen was shown to be located on intermediate filaments of cultured fibroblasts and to follow these filaments during various drug-induced rearrangements. As demonstrated by immunoprecipitation, antibody gel overlay and immunoblotting of two-dimensional polyacrylamide gels, VIC-C2 recognized two different antigens in extracts of THP-1 cells: one of Mr = 43 000 and pI = 7, the other of Mr = 57 000. In extracts from various cultured fibroblast cells only the 57 000 Mr antigen was detected. This 57 000 Mr protein was identified as vimentin by immunoblotting of rat glioma C6 cytoskeletons on two-dimensional gels. When vimentin was digested with chymotrypsin, only fragments containing parts of both helical rod pieces and the connecting non-helical spacer-region were strongly antigenic, whereas the helical rods alone were only weakly crossreactive. Moreover, immunoprecipitation revealed that VIC-C2 preferentially reacted with native compared to denatured vimentin.

Antibodies, Monoclonal↗

PubMed1985

Identification and spatial arrangement of high molecular weight proteins (Mr 300 000-330 000) co-assembling with microtubules from a cultured cell line (rat glioma C6).

The distribution of three high molecular weight proteins, MAP-1 (Mr 330 000), MAP-2 (Mr 300 000) and plectin (Mr 300 000) in various fractions obtained in cycles of temperature-dependent polymerization/depolymerization of microtubules from rat glioma C6 cells was studied. Using gel electrophoresis and immunoautoradiography/immunoblotting all three proteins were found to codistribute only partially with tubulin because considerable parts remained in the cold-insoluble fractions. Moreover, the proteins, particularly MAPs, were proteolytically degraded during cycling. By contrast, when microtubules were polymerized with taxol after isotonic cell lysis a considerable enrichment of MAP-1 and MAP-2 was achieved; again, plectin co-distributed only partially. In this procedure too, MAPs, especially MAP-2, were found to be highly subject to proteolysis, unless free Ca2+-ions were rigorously avoided. Proteolytic fragments generated from MAP-2 were of similar size independent of whether temperature- or taxol-dependent polymerization procedures were used, suggesting the occurrence of a MAP-2-specific protease. When the spatial arrangement of the high Mr proteins on taxol-polymerized C6 cell microtubules was directly visualized using gold-immunoelectron microscopy, a periodical, apparently helical, decoration of microtubules was found for MAP-1 and MAP-2; plectin was irregularly arrayed. A predominantly helical arrangement of both MAPs was demonstrated also for microtubules reconstituted from mammalian brain.

Alkaloids↗

PubMed1985

Morphological integrity of single adult cardiac myocytes isolated by collagenase treatment: immunolocalization of tubulin, microtubule-associated proteins 1 and 2, plectin, vimentin, and vinculin.

Single cardiac myocytes were isolated from hearts of 9 to 12-week-old rats by means of collagenase (100 U/ml). After assessment of their functional integrity they were processed for immunofluorescence microscopy of the cytoskeletal proteins tubulin, microtubule-associated proteins 1 and 2 (MAP-1 and MAP-2), plectin, vimentin, and vinculin. Antibodies to tubulin decorated a delicate filamentous network that apparently was unrelated to any sarcomeric organization. The distribution of MAP-1 and MAP-2 was strikingly different from that of tubulin, as both antigens were confined to Z-line structures. These structures were also prominently stained by affinity-purified antibodies to plectin and a monoclonal antibody to vimentin. Co-distribution of plectin and vimentin was also observed at the former intercalated disk region of the heart cell. Anti-vinculin antibodies decorated an intricate meshwork consisting of delicate filaments with predominantly irregular orientation and occasional assembly into whorls. These immunolocalization data indicate that the cell shape and cytoskeletal architecture characteristic of cardiac myocytes in tissues is maintained in single isolated cells. Furthermore, intermediate filaments rather than microtubules seem to be instrumental in the preservation of cell morphology.

Animals↗

PubMed1985

Expression of microtubule-associated proteins, MAP-1 and MAP-2, in human neuroblastomas and differential diagnosis of immature neuroblasts.

The expression of microtubule-associated proteins, MAP-1 and MAP-2, was studied in human neuroblastomas at various developmental stages using the immuno-alkaline-phosphatase technique and immunofluorescence microscopy. Of 15 cases examined, including grade I, grade II, and grade III neuroblastomas (M. Hughes, H. B. Marsden, and M. K. Palmer, Cancer 34:1706, 1974), rabbit antibodies raised against individual MAP-1 and MAP-2 from mammalian brain showed strong reactions with the whole spectrum of tumor cells including the immature small neuroblasts, partially mature neuroblasts, neurofibrils, and ganglion cells. Antibodies to alpha- and beta-tubulin showed similar staining patterns. In contrast, antibodies to the Mr = 200.000 neurofilament protein were reactive only with the mature and partially mature tumor cells, as well as with neurofibrils, but not with immature "round-cell" neuroblasts. Other types of round-cell tumors examined, including several cases of Ewing's sarcoma, undifferentiated rhabdomyosarcoma, and malignant lymphoma, showed no reaction with antibodies to MAP-1 and MAP-2. These tumors were reactive, however, with antibodies to various other tumor-specific as well as nonspecific antigens. It is concluded that antibodies to neuronal MAPs provide a valuable new tool for the differential diagnosis of neuroblastomas.

Adolescent↗

PubMed1984

Structural homology of microtubule-associated proteins 1 and 2 demonstrated by peptide mapping and immunoreactivity.

The major high molecular weight microtubule-associated polypeptides from hog brain (MAP-1 and MAP-2) were compared by one- and two-dimensional peptide mapping under varied conditions and by immunological techniques. Partial digestion of MAP-1 and MAP-2 with Staphylococcus aureus V8 protease and analysis in one dimension gave rise to very similar peptide maps independent of whether 125I-, 3H-, or 32P-labeled proteins were used. One-dimensional cleavage patterns of significant similarity were also obtained by partial digestion of MAP-1 and MAP-2 using trypsin or chymotrypsin. Furthermore, a pronounced similarity, although clear nonidentity, of MAP-1 and MAP-2 was also revealed after exhaustive digestion of 125I-labeled proteins with S. aureus V8 protease or trypsin followed by analysis of peptides in two dimensions. For immunological comparison, antisera were used that had been raised in rabbits using electrophoretically purified MAP-1 and MAP-2 components as immunogens. As determined by immunoprecipitation, the antiserum raised to MAP-1 was equally reactive with MAP-1 and MAP-2 components, whereas the antiserum to MAP-2 reacted primarily with MAP-2. Indicating the presence of common as well as unique antigenic determinants on MAP-1 and MAP-2, these results, therefore, were in agreement with the peptide mapping data. Implications of these results for biosynthetic mechanisms as well as differential distribution and functions of MAPs in cells are discussed.

Animals↗

PubMed1984

Widespread occurrence of polypeptides related to neurotubule-associated proteins (MAP-1 and MAP-2) in non-neuronal cells and tissues.

The occurrence and cellular localization of polypeptides related to hog brain microtubule-associated proteins 1 and 2 (MAP-1 and MAP-2) in non-neuronal cell lines of various species and types, and in several tissues from rat was studied. When insoluble cell fractions were prepared by incubation of isotonic cell extracts with 20 microM taxol, polypeptides co-migrating with MAP-1 and MAP-2 upon gel electrophoresis were observed in virtually all cases examined. Immunoblotting of preparations from 3T6, CHO, HeLa and N2A cells, as well as pituitary, heart, testis and liver revealed immuno-reactivity with antibodies to neuronal MAP-1 for polypeptides co-migrating with MAP-1 in all cases, except for HeLa cells and liver. With similar preparations, antibodies raised to neuronal MAP-2 were barely reactive with bands of the MAP-2 size except for N2A cells and pituitary gland. In all cases of non-neuronal cells and tissues, major cross-reactive bands, however, were of mol. wt. lower than that of MAP-2, indicating, most likely, proteolytic breakdown of MAP-2 during cell fractionation. As shown by double immunofluorescence microscopy of various cultured cell lines using affinity-purified antibodies to MAPs, and monoclonal antibodies to tubulin, MAP-1-as well as MAP-2-related antigens were generally, but not exclusively, associated with typical microtubule structures of the cytoplasm, spindle, midbody and primary cilia. Antigens related to both MAPs were also localized in frozen sections of rat trachea, testis, pituitary, kidney and cardiac and skeletal muscle.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1984

Identification of plectin in different human cell types and immunolocalization at epithelial basal cell surface membranes.

The occurrence of plectin in various human tissues and cell lines was investigated using immunofluorescence microscopy and antibody gel overlay/immunoblotting techniques. Plectin was identified in all tissues and cell lines tested, namely placenta, kidney, cornea, foreskin and eyelid skin, skin fibroblasts, monocytes, keratinocytes and HeLa cells. In frozen sections of cornea and skin, plectin was found to be enriched at epithelial basal cell surface membranes. Consequently, antibodies to plectin could serve as a tool in the classification of mechanobullous diseases.

Cell Line↗