Search PubMed⌕ Search

Biomedical subjects

M L Shelanski

Publications and source records attributed to M L Shelanski.

At least 19 recordsLinked to original sources
PubMed1992

Primary structure of high molecular weight tau present in the peripheral nervous system.

The tau proteins are a family of brain microtubule binding proteins that are required during axonal outgrowth and are found in neurofibrillary tangles in Alzheimer disease. A protein of higher molecular weight, immunologically related to tau, is expressed in the adult peripheral system and in cultured neuronal cell lines of neural crest origin. The predicted amino acid sequence of the high molecular weight tau from N115 cells has been determined from the sequence of its 2340-base-pair cDNA. High molecular weight tau contains an open reading frame encoding 733 amino acid residues. It contains sequences homologous to those present in the N-, middle, and C-terminal domains of adult brain tau proteins, including four homologous repeats, which are the tubulin binding sites, and an amino acid stretch, which is present only in the N-terminal domain of the mature brain variants. The middle region contains a previously unidentified nonhomologous stretch of 237 amino acid residues as well as a domain of 66 residues homologous to exon 6 of the bovine gene that is absent in all bovine, rat, and mouse tau cDNAs sequenced so far. A cDNA probe specific to the nonhomologous tau insert hybridizes to the 8- to 9-kilobase tau mRNA in N115 cells but not to the 6-kilobase tau mRNA in brain. Probes for the domains common to brain tau isoforms hybridize to both messages. The sequence of high molecular weight tau protein also suggests that it, like low molecular weight tau, is an elongated hydrophilic molecule. This cDNA should allow us to study the role of the domains specific to these tau forms in the specialization of the peripheral nervous system and for study of their expression in normal and pathological states.

Amino Acid Sequence↗
PubMed1992

Astrocyte process growth induction by actin breakdown.

cAMP analogues such as dibutyryl cAMP (dBcAMP) have been shown to induce the formation of processes in cultured primary astrocytes. We observe that the processes form by elongation as well as the previously reported retraction of cytoplasm around cytoskeletal elements. The most prominent cytoskeletal change that occurs in response to dBcAMP is a rearrangement of actin filaments characterized by a loss of cortical F-actin staining and the appearance of actin filament staining at the tips of the processes. If cortical actin filaments are disrupted with dihydrocytochalasin B, processes form that are similar to those induced by dBcAMP suggesting that the disruption of the cortical actin network is the pivotal step in process formation. Reorganization of the actin filament network in response to cAMP is accompanied by a decrease in phosphate incorporation into the regulatory light chain of myosin (MLC). Two selective inhibitors of MLC kinase (MLCK), ML-9 and KT5926, as well as a calmodulin antagonist (W7), which would also inhibit MLCK activation, all induce astrocytic process growth implicating MLCK as a control point in process initiation. We also found that dBcAMP and ML-9 both cause a decrease in the phosphate content of actin depolymerizing factor, suggesting that this protein and myosin light chain are the effectors of actin cytoskeleton reorganization and process growth.

Actin Cytoskeleton↗
PubMed1992

Neurite outgrowth in peripherin-depleted PC12 cells.

Peripherin is the major neuronal intermediate filament (IF) protein in PC12 cells and both its synthesis and amount increase during nerve growth factor (NGF) promoted neuronal differentiation. To address the question of the biological function of peripherin in neurite initiation we have used an antisense oligonucleotide complementary to the 5' region of peripherin mRNA to specifically inhibit its transcription. The oligonucleotide blocks both the synthesis of peripherin and its increase in response to NGF. Peripherin was found to be a stable protein with a cellular half-life of approximately 7 d. 6 wk of incubation with the oligonucleotide decreases peripherin to 11% of the level in naive control cells and to 3% of that in NGF-treated control cells. Despite the depletion, NGF elicits apparently normal neurite outgrowth from the oligonucleotide-treated cells. As evaluated by EM, there are few IFs in these cells, either in the cell bodies or neurites. There is no compensatory increase in NF-M, NF-L, or vimentin levels as a result of the inhibition of peripherin synthesis. These findings suggest that peripherin is not required for neurite formation, but is necessary for the formation of a cellular IF network which could be involved in process stability. They also demonstrate the utility of antisense oligonucleotides for the study of proteins with long half-lives.

Animals↗
PubMed1991

High molecular weight tau: preferential localization in the peripheral nervous system.

Using epitope mapping we have demonstrated that a high molecular weight protein (Mr approximately 115 x 10(3)) present in brain and spinal cord is a member of the tau family of microtubule-associated proteins. Antibodies directed against the amino-terminal, middle and carboxyl-terminal portions of tau recognize this protein. A limited survey of neuronal tissues has shown that this high molecular weight tau protein is present in brain, spinal cord, dorsal root ganglia, dorsal and ventral roots and peripheral nerves. High molecular weight tau protein is expressed at higher levels in spinal cord than in brain and is the only form of tau detected in the adult peripheral nervous system.

Animals↗
PubMed1990

Distribution of the protease inhibitor alpha 1-antichymotrypsin in cerebral and systemic amyloid.

We performed immunocytochemical staining to study the distribution of serum protease inhibitors in cerebral and systemic amyloid deposits. In beta-protein amyloid deposits in Alzheimer's disease, Down's syndrome, age-related cerebral amyloidosis, sporadic cerebral amyloid angiopathy and hereditary cerebral hemorrhage with amyloidosis of Dutch origin, antibody to alpha 1-antichymotrypsin (ACT) stains senile plaques and vascular deposits. Immature plaques or preamyloid deposits, identified by their positive staining for beta-protein and negative staining for Congo red, which represents the earliest recognizable stages of amyloid deposition, are also labeled. We did not detect ACT in other chemically different forms of cerebral and systemic amyloid. None of the other inhibitors in this study, i.e. antithrombin III and alpha 2-macroglobulin, was detected in the amyloid deposits. Neurons and glial cells throughout the central nervous system in normal and amyloid-containing brains also bind ACT antibody. The results emphasize the close association of ACT with one type of cerebral amyloid (beta-amyloid diseases) as well as the failure to detect such an association in other chemically different forms of cerebral and systemic amyloids.

Amyloid↗
PubMed1979

Biochemical and immunological characterization of neurofilaments in experimental neurofibrillary degeneration induced by aluminum.

In order to identify the protein composition of 10 nm neuronal filaments, we prepared enriched fractions of rabbit spinal neurons undergoing experimental neurofilamentous degeneration induced by aluminum. Electron microscopy of the isolated perikarya showed well-preserved, large perinuclear masses of neurofilaments, which were not found in similarly isolated control perikarya. Comparison of these glial-free fractions by SDS-polyacrylamide gel electrophoresis revealed several-fold augmentation in the filament-enriched neurons of proteins migrating at 68,000 and 160,000 daltons, with an additional component at 200,000 daltons. Otherwise, the protein patterns were identical; no band was found at 51,000 daltons, the molecular weight assigned to the major proteins both of glial filaments and of a previously reported bovine brain filament preparation. An antiserum raised against the 160,000 dalton component of a modified bovine brain filament fraction produced specific and intense fluorescent staining of the aluminum-induced neurofilament bundles. Antibodies to the 51,000 dalton protein of brain filaments and to tubulin failed to stain the induced filaments. The results strongly support the hypothesis that both normal and aluminum-induced neuronal filaments are composed of 68,000, 160,000 and 200,000 dalton polypeptides and do not contain significant amounts of the 51,000 dalton filament protein. The likelihood of biochemical heterogeneity among organelles with similar morphology, namely the glial and neuronal filaments, is raised.

Aluminum↗
PubMed1979

Neurofilaments.

Explore the source record for details and available documents.

Actins↗
PubMed1978

Intermediate filaments in nervous tissues.

Intermediate filaments have been isolated from rabbit intradural spinal nerve roots by the axonal flotation method. This method was modified to avoid exposure of axons to low ionic strength medium. The purified filaments are morphologically 75-80 percent pure. The gel electrophoretogram shows four major bands migrating at 200,000, 145,000, 68,000, and 60,000 daltons, respectively. A similar preparation from rabbit brain shows four major polypeptides with mol wt of 200,000 145,000, 68,000, and 51,000 daltons. These results indicate that the neurofilament is composed of a triplet of polypepetides with mol wt of 200,000, 145,000, and 68,000 daltons. The 51,000-dalton band that appears in brain filament preparations as the major polypeptide seems to be of glial origin. The significance of the 60,000- dalton band in the nerve root filament preparation is unclear at this time. Antibodies raised against two of the triplet proteins isolated from calf brain localize by immunofluorescence to neurons in central and peripheral nerve. On the other hand, an antibody to the 51,000-dalton polypeptide gives only glial staining in the brain, and very weak peripheral nerve staining. Prolonged exposure of axons to low ionic strength medium solubilizes almost all of the triplet polypeptides, leaving behind only the 51,000- dalton component. This would indicate that the neurofilament is soluble at low ionic strength, whereas the glial filament is not. These results indicate that neurofilaments and glial filaments are composed of different polypeptides and have different solubility characteristics.

Animals↗
PubMed1977

Specific neural and adrenal medullary antigens detected by antisera to clonal PC12 pheochromocytoma cells.

Antisera were prepared against a clonal line (PC12) of rat pheochromocytoma cells that were grown in either the presence or the absence of nerve growth factor (NGF). PC12 cells respond to NGF by growing neurites and express many differentiated properties of adrenal chromaffin cells and sympathetic neurons. The antisera, after absorption with rat liver, kidney, spleen, and thymus, reacted in the micro-complement fixation and indirect immunofluorescence assays with PC12 cells (both NGF-treated and untreated), brain, adrenal medulla, and superior cervical ganglia, but not with a variety of other tissues. Cross-absorption of the antisera with brain and adrenal medulla indicated the presence of components that were specific to brain, adrenal medulla, and PC12 cells. Brain-specific and adrenal-medulla-specific antigens were also detected in other species, such as rabbit, cat, and mouse. No differences could be detected in the specificity of antisera raised against either NGF-treated or untreated PC12 cells. Indirect immunofluorescence on live cells also detected cell surface antigens on both cell bodies and processes. These results suggest that such antisera may be used to detect, localize, and identify specific neural and adrenal medullary antigens.

Adrenal Medulla↗
PubMed1977

Localization of bovine brain filament antibody on intermediate (100 A) filaments in guinea pig vascular endothelial cells and chick cardiac muscle cells.

Guinea pig vascular endothelial cells contain naturally occurring rings of intermediate filaments that completely encircle the nucleus. Indirect immunofluorescence staining showed that these perinuclear rings bound antibody prepared against protein from bovine brain 9-nm filaments. In endothelial cells grown in the presence of 1 muM demecolcine (Colcemid) the perinuclear ring "coils" into a juxtanuclear "cap". Throughout this process we could demonstrate staining of the intermediate filaments. Chick cardiac muscle cells in culture stained diffusely with the antibody. After treatment for 24 hr with 1 muM demecolcine the cardiac cells accumulated large bands of intermediate filaments. These bands stained intensely with the antibody. Our findings suggest that intermediate filaments in guinea pig endothelial cells and those induced in chick cardiac muscle cells are antigenically similar to bovine brain filaments. The staining of these filaments is not affected by treatment with demecolcine.

Animals↗
PubMed1976

Biochemistry of the filaments of brain.

Intermediate filaments-cytoplasmic filaments with a diameter of 8-9 nm-have been described in a large variety of cell types. In this study, the subunit protein of the neurofilament and the presumptive subunit of the astroglial filament are compared by immunological and biochemical methods and are found to be very similar. Strong crossreactions are also found between these proteins in a variety of mammalian species. These results suggest that the intermediate filaments may, like microtubules and microfilaments, represent a highly conserved and widely distributed fibrous protein system.

Animals↗