Assignment of human alpha-synuclein (SNCA) and beta-synuclein (SNCB) genes to chromosomes 4q21 and 5q35.
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Animals↗
Biomedical subjects
M G Spillantini
Publications and source records attributed to M G Spillantini.
Somatodendritic localization and hyperphosphorylation of tau protein in transgenic mice expressing the longest human brain tau isoform.
Microtubule-associated protein tau is the major constituent of the paired helical filament, the main fibrous component of the neurofibrillary lesions of Alzheimer's disease. Tau is an axonal phosphoprotein in normal adult brain. In Alzheimer's disease brain tau is hyperphosphorylated and is found not only in axons, but also in cell bodies and dendrites of affected nerve cells. We report the production and analysis of transgenic mice that express the longest human brain tau isoform under the control of the human Thy-1 promoter. As in Alzheimer's disease, transgenic human tau protein was present in nerve cell bodies, axons and dendrites; moreover, it was phosphorylated at sites that are hyperphosphorylated in paired helical filaments. We conclude that transgenic human tau protein showed pre-tangle changes similar to those that precede the full neurofibrillary pathology in Alzheimer's disease.
Alzheimer Disease↗
Tau protein in Alzheimer's disease.
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Alzheimer Disease↗
Characterisation of an antibody relevant to the neuropathology of Alzheimer disease.
The specificity of BR88, a polyclonal antibody we raised against amino acids 1-12 of beta/A4 (beta-amyloid protein), has been reexamined in view of a claim made that it cross-reacts with tau-protein, probably the sole component of the paired helical filaments of Alzheimer disease. We have used enzyme-linked immunosorbent assays, immunoblots, and immunohistochemistry, together with appropriate controls, to show that BR88 does not cross-react with tau. This contradicts the claim in question and confirms our previous findings in which BR88 was used to show that some tangle-bearing cells displayed the N-terminus of beta/A4 on their surfaces. The conclusions formed in that earlier work therefore remain intact.
Adult↗
Identification of two distinct synucleins from human brain.
Two abundant proteins of 140 and 134 amino acids were purified and sequenced from human brain. They were identified through their reactivity on immunoblots with a partially characterised monoclonal antibody that recognises tau protein in a phosphorylation-dependent manner. The 140 amino acid protein is identical with the precursor of the non-A beta component of Alzheimer's disease amyloid which in turn is highly homologous to synuclein from Torpedo electroplaques and rat brain. The 134 amino acid protein is the human homologue of bovine phosphoneuroprotein 14; it is 61% identical in sequence to the 140 amino acid protein. The previously unrecognised homology between these two proteins defines a family of human brain synucleins. We refer to the 140 and 134 amino acid proteins as alpha-synuclein and beta-synuclein, respectively. Both synucleins are expressed predominantly in brain, where they are concentrated in presynaptic nerve terminals.
Alzheimer Disease↗
Neutral endopeptidase (3.4.24.11) in plasma and synovial fluid of patients with rheumatoid arthritis. A marker of disease activity or a regulator of pain and inflammation?
In recent years the role of the peripheral nervous system has been focused on the pathogenesis of rheumatoid arthritis (RA). In particular, substance P (SP), released by the sensory terminals, has been demonstrated to be involved in cartilage breakdown [13]. The aim of our work was to study the levels of SP and its peptidases, neutral endopeptidase (3.4.24.11) (NEP) and angiotensin-converting enzyme (ACE), in the synovial fluid and plasma of 30 patients with RA and 14 patients with osteoarthritis (OA). ACE and NEP were determined with a fluorimetric assay and SP with a radioimmunoassay (RIA) method. ACE levels were normal in the plasma of patients with RA and OA (6.1 +/- 1.9 and 6.7 +/- 1.4 pmol/ml/min, respectively); we found no differences in the values, of ACE between RA and OA synovial fluid (5.7 +/- 4.2 and 5.5 +/- 4.1 pmol/ml/min, respectively). NEP levels were significantly increased in plasma (139.3 +/- 36 pmol/ml/min) and synovial fluid (133.8 +/- 32 pmol/ml/min) of patients with RA when compared to patients with OA (73.4 +/- 22 in plasma and 15.2 +/- 10.8 pmol/ml/min in synovial fluid) and healthy controls (89.7 +/- 14 pmol/ml/min in plasma). In synovial fluid, SP was significantly higher in RA patients (43.1 +/- 16.6 pg/ml) than in OA patients (12 +/- 13.1 pg/ml), while plasma levels did not show any difference (RA: 14.4 +/- 10.2; OA: 13.6 +/- 10.6; healthy subjects: 11.3 +/- 3.9 pg/ml). The only relationship detected in controls and in OA was among plasma NEP and ESR (P < 0.05) and synovial fluid NEP (P < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)
Adult↗
The synovium of transgenic arthritic mice expressing human tumor necrosis factor contains a high level of nerve growth factor.
We have recently reported that nerve growth factor (NGF) increases in the synovium of patients affected by rheumatoid arthritis and in the synovium of pharmacologically-induced arthritis in animal models. In the present study, we demonstrate that arthritic transgenic mice which carry and express the human TNF gene (Tg197) also express elevated levels of NGF, and that subcutaneous injection of NGF-antibodies attenuates the loss of body weight caused by the development of disease in these mice. Along with our previous findings, which show an increase in the level of NGF during the acute phase of other autoimmune diseases, these results suggest a role of NGF in these pathologies. The functional significance of NGF in rheumatoid arthritis (RA) is currently under study.
Animals↗
Cloning of a big tau microtubule-associated protein characteristic of the peripheral nervous system.
Microtubule-associated protein tau consists in brain of a series of isoforms of 48- to 67-kDa apparent molecular mass that are encoded by mRNAs of approximately 6 kilobases (kb) and that are generated from a single gene by alternative splicing. Previously, a tau-like protein of 110-kDa apparent molecular mass was described in peripheral ganglia and in peripheral neuronlike cell lines. We now report the cloning and sequencing of a rat cDNA encoding this big tau. The corresponding protein contains sequence identical to the longest of the previously cloned small tau isoforms but with an additional 254 amino acid insert in the amino-terminal half. Big tau is produced from an 8-kb mRNA generated by alternative splicing from the same gene that encodes small tau. Production of big tau from the cloned sequence gives a protein of 110-kDa apparent molecular mass that aligns on SDS/PAGE with big tau protein extracted from peripheral ganglia. RNA blots show that in peripheral ganglia from adult rats only the 8-kb mRNA band corresponding to big tau is found, whereas in ganglia from newborn rats both 6- and 8-kb tau mRNA bands are found. In tissues from the central nervous system only the 6-kb mRNA band can be detected. Big tau protein is therefore produced specifically in the peripheral nervous system, and it will be interesting to see whether further molecular differences between the two major divisions of the vertebrate nervous system will be discovered.
Amino Acid Sequence↗
Tau proteins of Alzheimer paired helical filaments: abnormal phosphorylation of all six brain isoforms.
Preparations of dispersed paired helical filaments (PHFs) from the brains of Alzheimer's disease and Down's syndrome patients display on gels three principal bands corresponding to abnormally modified forms of the microtubule-associated protein tau. Interpretation of the pattern is difficult because there are six tau isoforms in normal brain and phosphorylation changes their mobility. By enzymatic dephosphorylation at high temperature, we have shifted the three abnormal bands obtained from dispersed PHFs to align with the six nonphosphorylated tau isoforms. By using antibodies specific for some of the inserts that distinguish the various isoforms and label PHFs, we have established a correspondence between PHFs, abnormal bands, and isoforms. This identification of isoforms is a necessary step in unravelling the molecular pathogenesis of PHFs.
Aged↗
The human glutamate receptor cDNA GluR1: cloning, sequencing, expression and localization to chromosome 5.
The rat glutamate receptor is a 907 amino acid transmembrane protein. Using the rat GluR1 cDNA as a probe, we have isolated cDNA clones from a human hippocampal cDNA library. Sequence of a full length cDNA clone revealed 98.2% and 89.4% identity to the rat sequence at the amino acid and nucleotide levels respectively. The human cDNA clone detected an RNA transcript in human cerebral cortex, hippocampus and cerebellum, similar to that seen in rat. In situ hybridization experiments showed that human GluR1 mRNA is present in granule and pyramidal cells in the hippocampal formation and that there is no apparent difference of distribution between control patient and patient with Alzheimer's disease. Dot blot analysis of flow-sorted human chromosomes showed that the GluR1 gene maps to chromosome 5.
Aged↗
Localization of the Alz-50 epitope in recombinant human microtubule-associated protein tau.
Alz-50 is a monoclonal antibody that stains the neurofibrillary pathology of Alzheimer's disease, as well as apparently normal nerve cells that are at risk of developing neurofibrillary tangles. On immunoblots it recognizes microtubule-associated protein tau and proteins of 60-68 kDa that are associated with Alzheimer's disease. We have used recombinant tau proteins expressed in E. coli to map the Alz-50 epitope to amino-terminal residues 2-10, a region common to all known human tau isoforms. A direct correspondence between immunoblots and histological staining was established by the abolition of Alz-50 staining following adsorption with recombinant tau proteins retaining amino-terminal sequences. This suggests that tau pathology represents an early event in the development of the neurofibrillary pathology of Alzheimer's disease.
Aged↗
Tau proteins and neurofibrillary degeneration.
The paired helical filament is the major fibrous component of neurofibrillary pathology in Alzheimer's disease. Over the last three years evidence has accumulated that the microtubule-associated protein tau forms an important, if not the sole, constituent of the paired helical filament. Tau protein in normal brain is bound to axonal microtubules by a tandem repeat region. In Alzheimer's disease a proportion of tau protein becomes abnormally phosphorylated and is no longer associated with axonal microtubules but instead accumulates in paired helical filaments throughout affected nerve cells. The tandem repeat region contributes substantially to the structural core of the paired helical filament, around which the amino-terminal half of the molecule forms a disordered coat.
Alzheimer Disease↗
Developmentally regulated and tissue specific expression of mRNAs encoding the two alternative forms of the LIM domain oncogene rhombotin: evidence for thymus expression.
The T-cell oncogene rhombotin was first identified as a gene near a chromosomal translocation breakpoint in a human T-cell tumour and represents the first example of an oncogene carrying the duplicated cysteine-rich regions (CRR or LIM domains). Transgenic expression of a reporter gene under the control of one of the rhombotin gene promoters subsequently showed high levels of expression in the developing brain. These disparate sites of transcriptional activity suggested that the gene may have been activated de novo specifically in the T cell tumour via the translocation. Here, we assess this possibility by analysing rhombotin gene expression in mouse development by in situ hybridization of whole embryos, Northern filter hybridization, and a sensitive semiquantitative PCR method. The results show that the central nervous system is the major site of rhombotin mRNA production. Low level expression does, however, occur in other tissues including thymus. Furthermore, both promoters are active and differentially regulated during mouse embryogenesis in both brain and thymus. In subregions of the adult brain, different levels of rhombotin activity can be observed, with evidence for regional variation in promoter usage. A detailed analysis of mouse and human T-cell differentiation suggests that fluctuating promoter activities are related to a general T-cell differentiation process rather than to the differentiation of functionally distinct subsets of T-cells. These data suggest that the transforming activity of rhombotin in the T-cell with the chromosomal translocation was not due to de novo transcriptional activation, but rather to a quantitative or qualitative change in expression levels of this CRR-containing oncogene after chromosomal translocation.
Animals↗
Characterization of endopeptidase 3.4.24.11 ("enkephalinase") activity in human plasma and cerebrospinal fluid.
The presence of the neutral metallo-endopeptidase 3.4.24.11 ("enkephalinase") activity was investigated by fluorimetric assay in human body fluids. Although the enzyme was previously known to occur exclusively in membrane bound form in the human or animal central nervous system, its activity was detected in human cerebrospinal fluid (CSF), plasma and amniotic fluid. Although the endopeptidase 3.4.24.11 activity found in human body fluids has properties closely related to the membrane bound enzyme such as affinity constant for the inhibitors, optimal pH and Km for the substrate, the Vmax values were in CSF, plasma and amniotic fluid, respectively, 2 x 10(3), 10(2), 10 lower than mouse brain homogenate.
Adult↗
Segmental and developmental regulation of a presumptive T-cell oncogene in the central nervous system.
Although most proto-oncogenes such as c-myc are involved in cell proliferation, being expressed in a wide range of tissues as well as in progenitors of transformed cells, others may normally function in cellular differentiation. We now report on a gene on human chromosome 11, at the junction of the T-cell tumour-associated chromosomal translocation t(11; 14) (p15; q11) and known as the 11p15 gene or Ttg, which is believed to be involved in the pathogenesis of the tumour. It has two transcriptional promoters (both retained by the translocated allele) and is expressed in tumour cells with neuro-endocrine properties, suggesting that normal expression may occur in nerve cells. Using fusion constructs of one 11p15 promoter and lacZ in transgenic mice, we found that the gene is expressed in a segment-specific manner in rhombomeres of the developing mouse hind-brain. During subsequent development, the gene is more widely expressed, again in precisely defined regional patterns, but in post-mitotic neurons confined to the central nervous system. Thus, this presumptive T-cell oncogene is both developmentally regulated and segmentally restricted in a tissue different from that in which the original tumour arose.
Animals↗
Molecular neuropathology of Alzheimer's disease: in situ hybridization studies.
1. Abundant senile plaques and neurofibrillary tangles in certain brain regions constitute the major neuropathological characteristics of Alzheimer's disease. Recent work has established that the amyloid beta protein, which is derived from a large precursor, constitutes the major constituent of plaque amyloid, whereas the microtubule-associated protein tau is a component of the paired helical filament, the major constituent of neurofibrillary tangles. 2. Multiple isoforms of amyloid beta protein precursor and tau protein are produced from a single gene through alternative RNA splicing. By Northern blotting amyloid beta protein precursor transcripts are found throughout central and peripheral tissues, whereas tau protein transcripts are only found in the nervous system. 3. In the central nervous system the cellular localization of amyloid beta protein precursor and tau protein transcripts is neuronal. The cells affected in Alzheimer's disease patients produce both types of transcripts; however, the various transcripts are also found in cells not affected in the course of the disease. At present, there exists no evidence to suggest that an overproduction of amyloid beta protein precursor or tau protein is the reason for plaque and tangle formation. The formation of the latter probably results from posttranslational events.
Alzheimer Disease↗
Different configurational states of beta-amyloid and their distributions relative to plaques and tangles in Alzheimer disease.
Antibodies have been raised against synthetic peptides corresponding to different parts of the beta-amyloid sequence. These antibodies stain different kinds of amyloid distributions in the hippocampal formation in Alzheimer disease, suggesting the existence of different states of aggregation and/or folding of beta-amyloid molecules. An antibody directed against the middle region of beta-amyloid stained mostly amyloid plaques without cores, whereas an antibody directed against the carboxyl-terminal region of beta-amyloid stained only amyloid plaques with cores. An antiserum directed against the amino terminus of beta-amyloid stained numerous tangle-bearing cells and bodies, as well as the neuritic component of plaques and neuropil threads. These antibodies, in conjunction with anti-tau antibodies, were used to demonstrate a close spatial relationship between amyloid deposits and neurofibrillary tangles.
Alzheimer Disease↗
Topographical relationship between beta-amyloid and tau protein epitopes in tangle-bearing cells in Alzheimer disease.
Double-labeling immunohistochemistry was used to investigate the topographical relationship between beta-amyloid and tau protein epitopes present in cells bearing neurofibrillary tangles found in the hippocampal formation of patients with Alzheimer disease. An antiserum raised against the amino terminus of beta-amyloid stained numerous tangle-bearing cells and other bodies ("extracellular tangles"), but double labeling showed that the beta-amyloid staining is invariably peripheral to that of the tau-positive tangle proper. This and other results suggest that the extracellular amyloid plaques and the intracellular neurofibrillary tangles are biochemically distinct but may result from related pathological events that originate at the level of the nerve cell and lead to its degeneration.
Alzheimer Disease↗