Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0027819 (neuroblastoma)
 27,800 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The gene defective in Huntington's disease encodes a protein, huntingtin, with unknown function. Antisera generated against three separate regions of huntingtin identified a single high molecular weight protein of approximately 320 kDa on immunoblots of human neuroblastoma extracts. The same protein species was detected in human and rat cortex synaptosomes and in sucrose density gradients of vesicle-enriched fractions, where huntingtin immunoreactivity overlapped with the distribution of vesicle membrane proteins (SV2, transferrin receptor, and synaptophysin). Immunohistochemistry in human and rat brain revealed widespread cytoplasmic labeling of huntingtin within neurons, particularly cell bodies and dendrites, rather than the more selective pattern of axon terminal labeling characteristic of many vesicle-associated proteins. At the ultrastructural level, immunoreactivity in cortical neurons was detected in the matrix of the cytoplasm and around the membranes of the vesicles. The ubiquitous cytoplasmic distribution of huntingtin in neurons and its association with vesicles suggest that huntingtin may have a role in vesicle trafficking.
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PMID:Huntingtin is a cytoplasmic protein associated with vesicles in human and rat brain neurons. 774 55

Huntington's disease is a progressive neurodegenerative disorder, which is caused by expansion of a polymorphic (CAG)n repeat in the coding region of the Huntington's disease gene. The function of huntingtin has not been elucidated so far. Accordingly, detailed subcellular localization studies remain useful. In an immunohistochemical study, we have reported huntingtin to be present in the cytoplasm of cells in the majority of the tissues studied. In addition, we detected a signal in the nucleus of cells in some tissues, including neuronal cells. We have further extended these studies in various mammalian cell lines, using a panel of (affinity-purified) polyclonal huntingtin antibodies in immunofluorescence, confocal laser scanning microscopy and biochemical subcellular fractionation studies. In mouse embryonic fibroblasts, human skin fibroblasts and in mouse neuroblastoma cells huntingtin was present in the cytoplasm. All five antibodies, directed against different parts of huntingtin, also showed a signal in the nucleus. This signal could be competed by the original antigen. The localization of huntingtin in both cytoplasm and nucleus, was confirmed by biochemical subcellular fractionation studies. However, in most other studies, a nuclear location for huntingtin has not been found. Our results suggest, however, that besides its function(s) in the cytoplasm, a nuclear function of huntingtin at some stages of differentiation or in some phases of the cell cycle may not be excluded.
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PMID:Subcellular localization of the Huntington's disease gene product in cell lines by immunofluorescence and biochemical subcellular fractionation. 884 26

Huntington's disease (HD) is a progressive neurodegenerative disorder caused by an expanding CAG repeat coding for polyglutamine in the huntingtin protein. Recent data have suggested the possibility that an N-terminal fragment of huntingtin may aggregate in neurons of patients with HD, both in the cytoplasm, forming dystrophic neurites, and in the nucleus, forming intranuclear neuronal inclusion bodies. An animal model of HD using the short N-terminal fragment of huntingtin has also been found to have intranuclear inclusions and this same fragment can aggregate in vitro . We have now developed a cell culture model demonstrating that N-terminal fragments of huntingtin with expanded glutamine repeats aggregate both in the cytoplasm and in the nucleus. Neuroblastoma cells transiently transfected with full-length huntingtin constructs with either a normal or expanded repeat had diffuse cytoplasmic localization of the protein. In contrast, cells transfected with truncated N-terminal fragments showed aggregation only if the glutamine repeat was expanded. The aggregates were often ubiquitinated. The shorter truncated product appeared to form more aggregates in the nucleus. Cells transfected with the expanded repeat construct but not the normal repeat construct showed enhanced toxicity to the apoptosis-inducing agent staurosporine. These data indicate that N-terminal truncated fragments of huntingtin with expanded glutamine repeats can aggregate in cells in culture and that this aggregation can be toxic to cells. This model will be useful for future experiments to test mechanisms of aggregation and toxicity and potentially for testing experimental therapeutic interventions.
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PMID:Truncated N-terminal fragments of huntingtin with expanded glutamine repeats form nuclear and cytoplasmic aggregates in cell culture. 953 81

Tissue transglutaminase is a calcium-dependent transamidating enzyme that has been postulated to play a role in the pathology of expanded CAG repeat disorders with polyglutamine expansions expressed within the affected proteins. Because intranuclear inclusions have recently been shown to be a common feature of many of these codon reiteration diseases, the nuclear localization and activity of tissue transglutaminase was examined. Subcellular fractionation of human neuroblastoma SH-SY5Y cells demonstrated that 93% of tissue transglutaminase is localized to the cytosol. Of the 7% found in the nucleus, 6% copurified with the chromatin-associated proteins, and the remaining 1% was in the nuclear matrix fraction. In situ transglutaminase activity was measured in the cytosolic and nuclear compartments of control cells, as well as cells treated with the calcium-mobilizing agent maitotoxin to increase endogenous tissue transglutaminase activity. These studies revealed that tissue transglutaminase was activated in the nucleus, a finding that was further supported by cytochemical analysis. Immunofluorescence studies revealed that nuclear proteins modified by transglutaminase exhibited a discrete punctate, as well as a diffuse staining pattern. Furthermore, different proteins were modified by transglutaminase in the nucleus compared with the cytosol. The results of these experiments clearly demonstrate localization of tissue transglutaminase in the nucleus that can be activated. These findings may have important implications in the formation of the insoluble nuclear inclusions, which are characteristic of codon reiteration diseases such as Huntington's disease and the spinocerebellar ataxias.
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PMID:Distinct nuclear localization and activity of tissue transglutaminase. 957 37

To gain insight into the pathogenic mechanisms of Huntington's disease (HD), we have developed a stable cellular model, using a neuroblastoma cell line in which the expression of full-length or truncated forms of wild-type and mutant huntingtin can be induced. While the wild-type forms have the expected cytoplasmic localization, the expression of mutant proteins leads to the formation of cytoplasmic and nuclear inclusions in a time- and polyglutamine length-dependent manner. The inclusions are ubiquitinated, appear more rapidly in cells expressing truncated forms of mutant huntingtin and are correlated with enhanced apoptosis. In lines expressing mutant full-length huntingtin, major characteristics present in Huntington's patients could be modelled. Selective processing of the mutant, but not the wild-type, full-length huntingtin was observed at late time points, with appearance of a breakdown product corresponding to a predicted caspase-3 cleavage product. A more truncated N-terminal fragment of huntingtin is also produced, that appears involved in building up cytoplasmic inclusions at early time points, and later on also nuclear inclusions. This fits with the finding that inclusions in the brain of HD patients are detected only using antibodies directed against epitopes very close to the polyglutamine stretch. This unique model should thus be useful to study the processing mechanism of mutant huntingtin, its role in the formation of intracellular aggregates and the effect of the latter on cellular physiology.
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PMID:A cellular model that recapitulates major pathogenic steps of Huntington's disease. 970 Jan 87

Huntingtin is a cytoplasmic protein of unknown function that associates with vesicle membranes and microtubules. Its protein interactions suggest that huntingtin has a role in endocytosis and organelle transport. In this study we sought to identify factors that regulate the transport of huntingtin in striatal neurons, which are the cells most affected in Huntington's disease. In clonal striatal cells derived from fusions of neuroblastoma and embryonic striatal neurons, huntingtin localization is diffuse and slightly punctate in the cytoplasm. When these neurons were differentiated by treatment with forskolin, huntingtin redistributed to perinuclear regions, discrete puncta along plasma membranes, and branch points and terminal growth cones in neurites. Huntingtin staining overlapped with clathrin, a coat protein involved in endocytosis. Immunoblot analysis of subcellular membrane fractions separated by differential centrifugation confirmed that huntingtin immunoreactivity in differentiated neurons markedly increased in membrane fractions enriched with clathrin and with huntingtin-interacting protein 1. Dopamine treatment altered the subcellular localization of huntingtin and increased its expression in clathrin-enriched membrane fractions. The dopamine-induced changes were blocked by the D1 antagonist SCH 23390 and were absent in a clonal cell line lacking D1 receptors. Results suggest that the transport of huntingtin and its co-expression in clathrin and huntingtin-interacting protein 1-enriched membranes is influenced by activation of adenylyl cyclase and stimulation of dopamine D1 receptors.
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PMID:Forskolin and dopamine D1 receptor activation increase huntingtin's association with endosomes in immortalized neuronal cells of striatal origin. 1036 4

Huntington's disease (HD) is a neurodegenerative disorder with a midlife onset. The disease is caused by expansion of a CAG (glutamine) repeat within the coding region of the HD gene. The molecular mechanism by which the mutated protein causes this disease is still unclear. To study the protein we have generated a set of rabbit polyclonal antibodies raised against different segments of the N-terminal, central and C-terminal parts of the protein. The polyclonal antibodies were affinity purified and characterized in ELISA and Western blotting experiments. All antibodies can react with mouse and human proteins. The specificity of these antibodies is underscored by their recognition of huntingtin with different repeat sizes in extracts prepared from patient-derived lymphoblasts. The antibodies were used in immunofluorescence experiments to study the subcellular localization of huntingtin in mouse neuroblastoma NIE-115 cells. The results indicate that most huntingtin is present in the cytoplasm, whereas a minor fraction is present in the nucleus. On differentiation of the NIE-115 cells in vitro, the subcellular distribution of huntingtin does not change significantly. These results suggest that full-length huntingtin with a normal repeat length can be detected in the nucleus of cycling and non-cycling cultured mammalian cells of neuronal origin. However, in HD autopsy brain the huntingtin-containing neuronal intranuclear inclusions can be detected only with antibodies raised against the N-terminus of huntingtin. Thus several forms of huntingtin display the propensity for nuclear localization, possibly with different functional consequences.
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PMID:Analysis of the subcellular localization of huntingtin with a set of rabbit polyclonal antibodies in cultured mammalian cells of neuronal origin: comparison with the distribution of huntingtin in Huntington's disease autopsy brain. 1043 6

The mitochondrial toxin 3-nitropropionic acid (3-NPA) causes neurodegeneration in the basal ganglia and neurological symptoms resembling Huntington's disease (HD) when applied to primates or rodents, and therefore might be used as an animal model for this disorder. For that reason, the molecular mechanisms involved in 3-NPA-induced neurodegeneration are of considerable interest. In our model, murine neuroblastoma cells (Neuro-2a) were treated with different doses of 3-NPA, and changes in gene expression were analyzed by means of mRNA differential display (DDRT-PCR). Using 18 primer combinations, we have identified a set of 33 candidate cDNAs deriving from 29 excised DDRT bands whose expression appeared to be changed in response to the 3-NPA insult (mostly elevated). DNA sequencing revealed that novel, as well as previously described genes, are included in this panel. Amongst the known cDNAs, the differential mRNA expression of the ribosomal proteins S6 and L40, of the protein kinase A (PKA) catalytic beta subunit and of the intercellular adhesion molecule ICAM-1 could be verified using Northern hybridization and RT-PCR, respectively. Furthermore, ICAM-1 expression could also be shown to increase at the protein level, which points to a possible function for this molecule in neuronal cells in the course of neurodegeneration. The results may prove useful in elucidating the multiple processes causing neurodegeneration subsequent to lesions by mitochondrial toxins and excitotoxins as well.
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PMID:Differentially displayed genes in neuroblastoma cells treated with a mitochondrial toxin: evidence for possible involvement of ICAM-1 in 3-nitropropionic acid-mediated neurodegeneration. 1081 91

The type I inositol 1,4,5-trisphosphate (IP(3)) receptor is selectively down-regulated in several neurodegenerative diseases, including Alzheimer's disease, Huntington's chorea, and ischemia, all conditions in which apoptotic neuronal loss occurs. In the present study, we used a neuronal cell line, human neuroblastoma SH-SY5Y cells, to investigate whether the levels of IP(3) receptor are changed during apoptosis in these cells. Following induction of apoptosis by staurosporine, the immunoreactivity of the type I IP(3) receptor in microsome preparations from SH-SY5Y cells was reduced within 2 h, with a further reduction during subsequent hours. Immunoblot analyses, using antibodies to poly(ADP-ribose) polymerase and spectrin breakdown products, revealed proteolysis of these caspase-3 substrates within 3 h, confirming that IP(3) receptor cleavage is an early consequence of apoptosis. In vitro incubation of SH-SY5Y microsomes or immunopurified IP(3) receptor from rat cerebellum with recombinant caspase-3 led to generation of immunoreactive breakdown products similar to those observed in intact cells, suggesting that the type I IP(3) receptor is a potential substrate for caspase-3. Preincubation of the neuroblastoma cells with the caspase-3 inhibitor Z-Asp-Glu-Val-Asp-fluoromethyl ketone prevented IP(3) receptor degradation. These results show that the type I IP(3) receptor is a substrate for caspase-3 in neuronal cells and indicate that apoptotic down-regulation of IP(3) receptor levels may contribute to the pathology of neurodegenerative conditions.
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PMID:Degradation of the type I inositol 1,4,5-trisphosphate receptor by caspase-3 in SH-SY5Y neuroblastoma cells undergoing apoptosis. 1103 74

Tissue transglutaminase (tTG) is a transamidating enzyme that is elevated in Huntington's disease (HD) brain and may be involved in the etiology of the disease. Further, there is evidence of impaired mitochondrial function in HD. Therefore, in this study, we examined the effects of mitochondrial dysfunction on the transamidating activity of tTG. Neuroblastoma SH-SY5Y cells stably overexpressing human tTG or mutated inactive tTG were treated with 3-nitropropionic acid (3-NP), an irreversible inhibitor of succinate dehydrogenase. 3-NP treatment of tTG-expressing cells resulted in a significant increase of TG activity in situ. In vitro measurements demonstrated that 3-NP had no direct effect on tTG activity. However, 3-NP treatment resulted in a significant decrease of the levels of GTP and ATP, two potent inhibitors of the transamidating activity of tTG. No significant changes in the intracellular levels of calcium were observed in 3-NP-treated cells. Treatment with 3-NP in combination with antioxidants significantly reduced the 3-NP-induced increase in in situ TG activity, demonstrating that oxidative stress is a contributing factor to the increase of TG activity. This study demonstrates for the first time that impairment of mitochondrial function significantly increases TG activity in situ, a finding that may have important relevance to the etiology of HD.
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PMID:Impaired mitochondrial function results in increased tissue transglutaminase activity in situ. 1103 84


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