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J Avila

Publications and source records attributed to J Avila.

At least 199 records · Page 11Linked to original sources
PubMed1990

Aluminum induces the in vitro aggregation of bovine brain cytoskeletal proteins.

The addition of aluminum to purified cytoskeletal proteins in vitro selectively induces the aggregation of highly phosphorylated proteins, such as the two larger neurofilament subunits (200 and 160 kDa) and the microtubule-associated proteins of the MAP-1 group (MAP-1A and MAP-1B). Other cytoskeletal proteins with a substantially lower phosphate content, such as the smaller neurofilament subunit (68 kDa) and tubulin, remain soluble, even in the presence of high aluminum concentrations. This suggests that aluminum interacts with phosphate groups in cytoskeletal proteins, causing their precipitation. The protein aggregates formed in the presence of aluminum are resistant to reagents such as urea and sodium dodecyl sulphate (SDS) which dissolve normal cytoskeletal polymers (neurofilaments and microtubules). These results favor the view that the neurotoxic effect of aluminum may be due primarily to the disorganization of the neuronal cytoskeleton which may occur subsequent to the precipitation of certain highly phosphorylated cytoskeletal proteins.

Aluminum↗
PubMed1990

Characterization of tau protein present in microtubules and paired helical filaments of Alzheimer's disease patient's brain.

Two proteins immunologically related to porcine tau protein are found in the brain of Alzheimer's disease patients. One is bound to microtubules and, after isolation by co-polymerization with tubulin, shows a size and tryptic peptide map, similar to the microtubule-associated tau protein, present in the brain of non-demented patients. The other tau-related protein is present as the major protein of a purified fraction of paired helical filaments. The paired helical filament-associated protein shows smaller molecular weight (33,000) than microtubule-associated tau; however, this 33,000 mol. wt protein reacts with a monospecific anti-tau antibody and with an antibody to a 19-amino acid peptide corresponding to amino acids 228-246 of human tau. Furthermore, the 33,000 mol. wt protein and the tau protein have similar tryptic peptide maps. These results suggest that the paired helical filament protein is a modified form of the microtubule-associated tau protein.

Aged↗
PubMed1990

Collagenous structures present in brain contain epitopes shared by collagen and microtubule-associated protein tau.

A novel type of collagenous fibers has been isolated from human brain and characterized by electron microscopy and optical diffraction. It was found that the morphology of the fibers is similar, but not identical, to that of skin collagen. Also, the collagenous fibers show some similarities with the paracrystals that could be assembled in vitro from purified microtubule-associated protein tau. Immunological analyses indicated the presence of epitopes in these collagenous fibers which react with antibodies against collagen and tau.

Alzheimer Disease↗
PubMed1990

Microtubule dynamics.

A combination of biochemical, structural, and morphological analyses during the last 2 decades has shown that the cytoplasm of a cell is not a disorganized mass of jelly but a highly structured cell compartment formed of a cytoskeleton, one of which principal components are the microtubules. More recently, studies have revealed that microtubule cytoskeleton is not only well organized but highly dynamic, and that microtubule dynamics may be responsible for several cell functions such as chromosome segregation, cell morphogenesis, or intracytoplasmic organization.

Animals↗
PubMed1990

Phosphorylation of microtubule proteins in rat brain at different developmental stages: comparison with that found in neuronal cultures.

The phosphorylation of rat brain microtubule protein on intracranial injection of labeled phosphate has been analyzed. The major microtubule protein components phosphorylated in vivo in rat brain are the high-molecular-weight microtubule-associated proteins (MAPs) MAP-1A, MAP-1B, and MAP-2. A slight phospholabeling of beta-tubulin, which corresponds to the phosphorylation of a minor neuronal beta-tubulin isotype, is also observed. Whereas MAP-1B, MAP-2, and beta-tubulin are phosphorylated in the brain of 5-day-old rat pups, when most neurons of the CNS are extending processes, MAP-1A phosphorylation is observed only after neuronal maturation takes place. The phosphorylation of MAP-1A, MAP-1B, and beta-tubulin may be due mainly to casein kinase II or a related enzyme, whereas MAP-2 appears to be modified by other enzymes such as the cyclic AMP-dependent protein kinase (protein kinase A) and the calcium/phospholipid-dependent protein kinase (protein kinase C). Microtubule protein phosphorylation has also been studied in neuronal cultures. In differentiated neuroblastoma cells, only MAP-1B and beta-tubulin are phosphorylated in a manner coupled to neurite outgrowth. In primary cultures of fetal rat brain neurons, the pattern of microtubule protein phosphorylation resembles that found in vivo in rat pup brain. As phosphorylated MAP-1A and MAP-1B are present mainly on assembled microtubules, whereas the phosphorylation of MAP-2 decreases its interaction with microtubules, a role can be suggested for the phosphorylation of these proteins in the regulation of microtubule assembly and disassembly during neuronal development.

Aging↗
PubMed1990

Abnormal meiotic spindles cause a cascade of defects during spermatogenesis in asp males of Drosophila.

Since spermatogenesis in Drosophila is a series of interconnected and interdependent steps and most of the spermatogenic events take place in the absence of transcription, failures in a given stage can give rise to a cascade of defects later on. The asp locus of Drosophila melanogaster codes for a non-tubulin component implicated in proper spindle structure and/or function (Ripoll et al. 1985). Homozygous asp males exhibit abnormal meiotic spindles giving rise to altered segregation of chromosomes and mitochondria and failures in cytokinesis. Postmeiotic spermatogenic stages of asp males show a series of alterations that we interpret as due to the previously occurring defective meiosis because meiotic spindles are the only microtubular structure altered in mutant testes. The most conspicuous alterations are: (i) variable size of nuclei and nebenkerns of early spermatids, which are also multinucleate instead of having single and uniformly sized nuclei; (ii) elongating spermatids in which abnormal-sized mitochondrial derivatives elongate alongside more than one axoneme; (iii) failures in the individualization process, where abnormal spermatids remain syncytial, and seem to be eliminated during the coiling stage.

Animals↗
PubMed1990

A subset of antibodies from the sera of patients with systemic lupus erythematosus react with vimentin and DNA.

Sera from patients with systemic lupus erythematosus were tested for the simultaneous presence of antibodies to intermediate filaments (vimentin) and to DNA, using radioimmunoassay and immunofluorescence techniques. Our results indicate that 3 of 17 sera tested contain an IgM population which recognizes an antigenic determinant common to vimentin and DNA by a solid phase immunoassay.

Adolescent↗
PubMed1989

Sodium butyrate induces major morphological changes in C6 glioma cells that are correlated with increased synthesis of a spectrin-like protein.

Butyrate induced flattening and development of cell processes in rat glioma (C6) cells and this change was correlated with an increase in the synthesis of a polypeptide doublet with an apparent molecular weight of about 200 kDa. Blot analysis revealed that at least one of these polypeptides was a spectrin-like protein. Indirect immunofluorescence studies with the spectrin antiserum indicated that the antigen was present in the cell bodies, and also in the cell processes. Thus fodrin may be one the major targets for the action of butyrate on C6 cells.

Autoradiography↗
PubMed1989

Altered levels of microtubule proteins in brains of Alzheimer's disease patients.

The amount of microtubule protein present in the total soluble protein from brains of Alzheimer's disease patients and from brains of non-Alzheimer age-matched controls, were determined by radioimmunoassay. No differences were found in the amount of tubulin or microtubule-associated protein MAP2 present in either group. However, the amount of tau protein or MAP1 from the brains of Alzheimer's disease patients was about half of that present in their control counterparts.

Alzheimer Disease↗
PubMed1989

Subcellular localization of iodinated thyroid tubulin.

Subcellular fractions enriched in mitochondria, plasma membranes, microsomes and Golgi apparatus were obtained from thyroid glands of rats injected with I125. Autoradiography of SDS-polyacrylamide gels revealed the presence of a number of radiolabelled proteins in each membrane fraction. One polypeptide, with the same electrophoretic mobility as brain tubulin, was found in all fractions except the plasma membranes and was immunoprecipitated with commercial anti-tubulin monoclonal antibodies. Hydrolysis of Asp-Pro linkages of I125 labelled tubulin with formic acid indicated that there were iodination sites in both the carboxy terminal one third and the amino terminal two thirds of the molecule. These results, together with the absence of iodinated tubulin from the cytosolic fraction, are consistent with the idea that a population of thyroid membrane tubulin is iodinated at multiple sites either just before or after insertion into intracellular membranes where it may act as an anchorage point for microtubule-membrane interactions.

Animals↗
PubMed1989

A discrete repeated sequence defines a tubulin binding domain on microtubule-associated protein tau.

The protein domain responsible for the interaction of tau with tubulin has been identified. Biophysical studies indicated that the synthetic peptide Val187-Gly204 (VRSKIG-STENLKHQPGGG) from the repetitive sequence on tau binds to two sites on the tubulin heterodimer and to one site on each of the microtubule-associated protein-interacting C-terminal tubulin peptides alpha(430-441) and beta(422-434). The binding data showed a relatively stronger interaction of Val187-Gly204 with beta(422-434) as compared to that with alpha(430-441). The interaction of this tau peptide with either alpha or beta tubulin peptides appears to be associated with conformational changes in both the tau and the tubulin peptides. The beta tubulin peptide also appears to induce a structural change of tau fragment Val218-Gly235. Interestingly, tau peptides Val187-Gly204 and Val218-Gly235 induced tubulin self-assembly in a cold-reversible fashion, and incorporated into the assembled polymers. The specificity of the interaction of the tau peptide was supported by the competition of tau protein for the interaction with the tubulin polymer. In addition, the tau peptide appears to contain the principal antigenic determinant(s) recognized by anti-idiotypic antibodies that react with the tubulin binding domains on microtubule-associated proteins. The present findings together with the demonstration of the presence of multiple sites for the binding of the alpha(430-441) and beta(422-434) tubulin fragments to tau, and the existence of repetitive sequences on tau, strongly support the hypothesis that the region of tau defined by the repetitive sequences is involved in its interaction with tubulin.

Amino Acid Sequence↗
PubMed1989

Association of casein kinase II with microtubules.

A magnesium-dependent heparin-inhibited protein kinase activity associated with brain microtubule preparations has been identified as casein kinase II using a monospecific polyclonal antibody. This enzyme appears enriched in cold-stable microtubule fractions. By immunofluorescence microscopy using an antiserum against casein kinase II, the in situ immunolabeling of some microtubule assays has been observed. Thus, mitotic spindles are stained by the anti-casein kinase II antibody in fibroblast cells. In neuroblastoma cells induced to differentiate, the labeling of microtubule arrays inside developing axon-like processes is also seen. These results support the view that casein kinase II can modulate cytoskeletal assembly and dynamics through phosphorylation of microtubule proteins.

Animals↗
PubMed1989

Quantitation of microtubule-associated protein MAP-1B in brain and other tissues.

1. The presence of microtubule-associated protein MAP-1B in all mammalian tissues tested, as well as in brain, has been demonstrated by immunoblotting using a monospecific polyclonal antibody. 2. The expression of brain MAP-1B is developmentally controlled, as it is less abundant in adult than in newborn rat brain, where it is a major microtubule assembly promoting factor. 3. The level of MAP-1B in tissues other than brain is lower than it is in brain; but the relative ratios of MAP-1B to tubulin are very similar in all tissues, thus differing from the observed for MAP-2 or tau. 4. The amount of MAP-1B in non-nervous tissues seems not to be under developmental control. 5. These results are consistent with a role for MAP-1B in the assembly of microtubules in most cells.

Adrenal Glands↗
PubMed1989

Rearrangement of microtubule associated protein parallels the morphological transformation of neurons from dorsal root ganglion.

In primary cultures of dorsal root ganglion cells from rat embryos, neurons undergo a morphological transformation from a bipolar to a differentiated pseudo-unipolar shape, resembling their developmental stages in vivo. Cells present in these cultures are characterized here by immunological criteria using monoclonal and polyclonal antibodies against microtubule associated proteins MAP1 and MAP2 and against tubulin. After development for seven days in culture, antibodies against microtubule associated proteins MAP1 brightly labeled cells with neuronal morphology and lightly stained cells with the shape of Schwann cells. In addition, an extended network of neuronal processes was labeled with this antibody. Anti-microtubule associated protein MAP2 stained only neurons and a more restricted network of neuronal processes. The compartmentalization of microtubule associated protein MAP2 during the maturation process was followed by double-labeling with antibodies to microtubule associated proteins MAP1 and MAP2. Initially, microtubule associated protein MAP2 was present in the cell body and the two processes of bipolar neurons. Subsequently, the labeling of both processes changed, depending on neuronal morphology. In neurons in which both processes were approaching one another, one of these neurites was stained predominantly with anti-microtubule associated protein MAP2. Finally, in pseudo, unipolar neurons, anti-microtubule associated protein MAP2 labeling was found in the cell body and excluded from the more distal processes.

Animals↗
PubMed1989

Characterization of proteins immunologically related to brain microtubule-associated protein MAP-1B in non-neural cells.

Brain microtubule-associated protein MAP-1 is composed of at least two polypeptides, MAP-1A and MAP-1B, which are among the main components of the neural cytoskeleton. Specific monoclonal and polyclonal antibodies against MAP-1B stain nuclei, mitotic spindles, centrosomes and the cytoplasmic microtubule network of different non-neural cells studied by immunofluorescence microscopy. It appears that these cells contain two proteins of 325K and 220K (K = 10(3) Mr), which are immunologically related to brain MAP-1B. The 325K protein, which is localized to the cytoplasmic microtubule network, the centrosome and the mitotic spindle, seems to be structurally related to the neural MAP-1B, as judged from their similar peptide maps and phosphorylation patterns. The 220K protein, which is localized to the nuclear matrix in interphase cells and to the mitotic spindle in dividing cells, has a proteolytic profile different from that of neural MAP-1B and is phosphorylated to a much lesser extent than the 325K protein. Both proteins bind tubulin in vitro, which suggests that they may participate in microtubule assembly in vivo; the 325K protein could perform such a role during the entire cell cycle, while the 220K protein could be implicated in the formation of the mitotic spindle.

Amino Acids↗
PubMed1989

Calcium binding properties of the beta tubulin subunit from chicken erythrocytes.

Taxol-stabilised erythrocyte microtubules assembled less readily than similarly prepared brain microtubules on adding 10(-4) M-10(-3) M concentrations of calcium at 2 degrees C. Scatchard plot analyses of the high affinity calcium binding sites showed that the erythrocyte tubulin contained only 0.9 high affinity binding sites per dimer compared to 1.4 binding sites per dimer for brain tubulin. Association constants, however, for calcium binding to both erythrocyte and brain tubulin were similar (3.0 x 10(-6) M and 2.1 x 10(-6) M). The beta-tubulin subunit appeared to be responsible for the lower calcium binding ability of erythrocyte tubulin as shown by a gel overlay assay with 45Ca. Strains-all, a dye that stains many calcium binding proteins blue, did not stain erythrocyte beta-tubulin or its chymotryptic C-terminal fragment blue as was the case for brain beta-tubulin and its chymotryptic C-terminal fragment. We suggest that the lower calcium binding ability of erythrocyte beta-tubulin may be implicated in the differential behaviour of erythrocyte microtubules.

Alkaloids↗
PubMed1988

Detection of tubulin-binding proteins by an overlay assay.

A blot overlay technique to detect tubulin-binding proteins has been developed. It involves the fractionation of the putative tubulin-binding proteins by gel electrophoresis, their transfer to nitrocellulose paper, and the incubation of the nitrocellulose sheet with purified tubulin. The proteins which bind tubulin are recognized by monoclonal anti-tubulin antibodies. Proteins which associate with microtubules through a direct interaction with tubulin are easily detected by this procedure, which has been tested for both known microtubule-associated proteins and newly reported tubulin-binding proteins.

Animals↗