Amyotrophic lateral sclerosis (ALS) is a fatal disease characterized by the premature loss of motor neurons. While the underlying cellular mechanisms of neuron degeneration are unknown, the cytoplasmic aggregation of several proteins is associated with sporadic and familial forms of the disease. Both wild-type and mutant forms of the RNA-binding proteins FUS and TDP-43 accumulate in cytoplasmic inclusions in the neurons of ALS patients. It is not known if these so-called proteinopathies are due to a loss of function or a gain of toxicity resulting from the formation of cytoplasmic aggregates. Here we present a model of FUS toxicity using the yeast Saccharomyces cerevisiae in which toxicity is associated with greater expression and accumulation of FUS in cytoplasmic aggregates. We find that FUS and TDP-43 have a high propensity for co-aggregation, unlike the aggregation patterns of several other aggregation-prone proteins. Moreover, the biophysical properties of FUS aggregates in yeast are distinctly different from many amyloidogenic proteins, suggesting they are not composed of amyloid.
Amyotrophic Lateral Sclerosis ; metabolism ; pathology ; Cell Proliferation ; drug effects ; Cytoplasm ; drug effects ; metabolism ; DNA-Binding Proteins ; genetics ; metabolism ; Detergents ; pharmacology ; Humans ; Kinetics ; Peptides ; metabolism ; Prions ; chemistry ; metabolism ; Protein Binding ; drug effects ; Protein Multimerization ; drug effects ; Protein Structure, Quaternary ; Protein Transport ; RNA-Binding Protein FUS ; chemistry ; genetics ; metabolism ; Saccharomyces cerevisiae ; cytology ; drug effects ; genetics ; metabolism ; Saccharomyces cerevisiae Proteins ; chemistry ; metabolism

DEDD is a member of the death-effector domain protein family. DEDD inhibits the Smad3 mediated transcriptional activity and participates in the regulation of apoptosis. In this study, how the death-effector domain of DEDD participates in the regulation of Smad3 activity and apoptosis has been further investigated. Immunoblotting, immunofluorescence and immunoprecipitation had been used to detect the effects of the full length DEDD and its two truncated mutants, N-DEDD and C-DEDD on Smad3 subcellular distribution, phosphorylation, and interaction between Smad4. The effects of the full length DEDD and its two truncated mutants on cell apoptosis and proliferation had also been explored by flow cytometry and MTT assay. It showed that DEDD and N-DEDD inhibit TGF-beta1 induced Smad3 nuclear translocation and the formation of Smad3-Samd4 complex. DEDD and its two mutants can induce cell apoptosis and inhibit cell proliferation. These results suggested that DEDD inhibits the activity of Smad3 through its death-effector domain. Both the two truncated mutants of DEDD participate in the regulation of apoptosis and cell proliferation.
Apoptosis ; drug effects ; Cell Proliferation ; drug effects ; DNA-Binding Proteins ; pharmacology ; Death Domain Receptor Signaling Adaptor Proteins ; pharmacology ; HEK293 Cells ; Hep G2 Cells ; Humans ; Phosphorylation ; drug effects ; Protein Binding ; Smad3 Protein ; metabolism ; Smad4 Protein ; metabolism

OBJECTIVETo investigate the possible mechanisms of nobiletin for anticancer by studying the inhibition effects of nobiletin on tubulin polymerization.

METHODIn vitro nobiletin was added into the tubulin polymerization-depolymerization system and the absorption values were recorded at 350 nm under 37 degrees C.

RESULTAs compared with controls, the absorption values in reaction system decreased significantly in nobiletin treatment groups. When nobiletin final concentrations in reaction system were 5.0, 7.5, 10.0 and 12.5 micromol x L(-1), the maximum absorption values were 0 130, 0.109, 0.086 and 0.071 with 16.7%, 30.1%, 44.9% and 54.5% of inhibition rate, respectively. The results suggested that nobiletin could inhibit tubulin polymerization.

CONCLUSIONThe inhibition effect of nobiletin on tubulin polymerization is the possible mechanism for anticancer.

Animals ; Flavones ; pharmacology ; Protein Binding ; drug effects ; Swine ; Tubulin ; chemistry ; metabolism ; Tubulin Modulators ; pharmacology

Brain ; Cholesterol ; chemistry ; Cytoskeleton ; drug effects ; Endothelial Cells ; cytology ; metabolism ; Hemolysin Proteins ; chemistry ; pharmacology ; Humans ; Phalloidine ; pharmacology ; Pseudopodia ; drug effects ; Stress Fibers ; drug effects ; rac1 GTP-Binding Protein ; metabolism ; rhoA GTP-Binding Protein ; metabolism

OBJECTIVETo investigate the effects of enriched environment and impoverished environment on the learning and memory ability of manganese-exposed mice and the mechanism.

METHODSForty female Kunming mice were randomly and equally divided into 4 group: control group (CG), standard environment and manganese exposure group (SEG), enriched environment and manganese exposure group (EEG), and impoverished environment and manganese exposure group (IEG). The mouse model of manganese poisoning was established by intraperitoneal injection of manganese chloride. The learning and memory ability was tested by Morris water maze. The expression of cAMP response element-binding protein (CREB) in area CA1 of the hippocampus was measured by immunohistochemistry.

RESULTSIn place navigation test, the SEG had a significantly longer escape latency than the CG (P < 0.05), and the EEG had a significantly shorter escape latency than the SEG (P < 0.05); there was no significant difference in escape latency between IEG and SEG (P > 0.05). In spatial probe test, the EEG had a significantly greater number of platform crossings than the SEG (P < 0.05), and the IEG had a significantly smaller number of platform crossings than the SEG (P < 0.05). The expression of CREB in area CA1 of the hippocampus was significantly lower in IEG and SEG than in CG (P < 0.05), and it was significantly higher in EEG than in SEG (P < 0.05).

CONCLUSIONIn the enriched environment, the learning and memory ability of manganese-exposed mice can be improved, which may be due to the increased expression of CREB in the hippocampus.

Animals ; Cyclic AMP Response Element-Binding Protein ; metabolism ; Disease Models, Animal ; Environment ; Female ; Hippocampus ; drug effects ; metabolism ; Learning ; drug effects ; Manganese Poisoning ; metabolism ; Memory ; drug effects ; Mice

OBJECTIVETo explore the influence of beta-elemene on the proliferation, migration and RhoA expression of hepatic stellate cells (HSC) induced by angiotensin II (ANG II).

METHODSHSC were incubated in vitro. Proliferation and migration of the HSC were induced by ANG II. The effect on the proliferation of HSC was determined by MTT colorimetry. The migration ability was detected by transwell chamber cultures. Total RNA was extracted by TRizol reagent and gene levels were determined by semi-quantitative RT-PCR. Protein levels were determined by Western blot.

RESULTSDifferent concentrations (from 1 to 10 micromol/L) of ANG II markedly promoted the growth of the HSC in a concentration dependent way (0 micromol/L ANG II, F = 112.640, P less than 0.01). 10, 8, 4 micromol/L ANGII significantly induced HSC migration, F = 117.496, P less than 0.01. Compared with the 4 micromol/L ANG II group, 10 mg/L, 5 mg/L, 2.5 mg/L beta-elemene markedly inhibited HSC proliferation and migration induced by 4 micromol/L ANG II (F values were 95.706 and 55.600 and P less than 0.01). 4 micromol/L ANG II markedly promoted the protein and mRNA expressions of RhoA in HSC. 10 mg/L, 5 mg/L and 2.5 mg/L beta-elemene notably inhibited the expressions of RhoA protein and mRNA (F values were 217.119 and 18.010).

CONCLUSIONANG II can significantly induce the proliferation and migration of HSC. Beta-elemene can inhibit the proliferation and migration of HSC induced by ANG II. The effects of beta-elemene are mediated through inhibiting the RhoA signal transduction pathway and are associated with RhoA.

Angiotensin II ; pharmacology ; Cell Movement ; drug effects ; Cell Proliferation ; drug effects ; Cells, Cultured ; Hepatic Stellate Cells ; cytology ; drug effects ; Humans ; Sesquiterpenes ; pharmacology ; rhoA GTP-Binding Protein ; metabolism

OBJECTIVETo investigate the effects of hydroquinone (HQ) on expression of ubiquitin-ligating enzyme Rad18 in human hepatic cells (L-02), and to explore the role and possible mechanism of Rad18 involved in toxicity of HQ to hepatic cells.

METHODSAfter L-02 hepatic cells were exposed to HQ with various concentrations (0, 5, 10, 20, 40, 80 and 160 micromol/L) for 24 h, cell survival rate was measured by MTT assay; DNA impairment was evaluated by single cell gel electrophoresis (SCGE); The expression levels of Rad18 mRNA and protein were detected by Real-time fluorescent quantitative polymerase chain reaction (QPCR) technique and Western blot method respectively.

RESULTSHQ with concentration from 0 to 80 micromol/L had little effect on survival rate of L-02 (P > 0.05); Whereas the survival rate in the group of 160 micromol/L was significantly lower than in the control with the significant difference (P < 0.01) after treated with HQ for 24 h; The higher dose of HQ presented, the more degrees of olive tail moment (OTM) were produced and a dose-dependent relationship was shown. HQ in a low concentration (0 to approximately 40 micromol/L) could induce increase in the expression of Rad18 mRNA and protein which was in proportion to the increment of HQ concentration; the expression of Rad18 mRNA was enhanced increasingly, while the expression of Rad18 protein unchanged basically once the concentration of HQ exceeded 40 micromol/L; Besides, there was a positive correlation between OTM and the expression level of Rad18 mRNA (r = 0.919, P < 0.01).

CONCLUSIONHQ could regulate up the expression of Rad18 in L-02 hepatic cells.

Cell Survival ; drug effects ; Cells, Cultured ; DNA Damage ; drug effects ; DNA-Binding Proteins ; metabolism ; Hepatocytes ; drug effects ; enzymology ; Humans ; Hydroquinones ; toxicity ; Ubiquitin-Protein Ligases

Dalbergiae Odoriferae Lignum as a traditional Chinese medicine (TCM) has been widely used for promoting blood circulation and removing blood stasis. Flavonoid compounds are main chemical constituents of Dalbergiae Odoriferae Lignum, which exert anti-inflammatory property. However, the underlying anti-inflammatory mechanisms of flavonoid compounds are incompletely understood. It has been reported that isoliquiritigenin, liquiritigenin, naringenin and butein possess anti-inflammatory property. The purpose of this study is to illuminate the anti-inflammatory mechanism of flavonoid compounds based on the protein interaction network (PIN) analysis on molecular network level. 130 targets of the main medicinal ingredients of flavonoid compounds were gained though database retrieval. A protein interaction network of flavonoid compounds was constructed with 589 nodes and 216 interactions. By a graph theoretic clustering algorithm Molecular Complex Detection (MCODE), 26 modules were identified and analyzed by Gene ontology (GO) enrichment. Two modules were associated with anti-inflammatory actions. The most interesting finding of this study was that the anti-inflammatory effect of flavonoid compounds may be partly attributable to inhibite FOS, PTGS2 expression, inhibite of IL-1beta release, and block the MAPK pathway and toll-like receptor pathway.
Anti-Inflammatory Agents ; pharmacology ; Dalbergia ; chemistry ; Drugs, Chinese Herbal ; pharmacology ; Flavonoids ; pharmacology ; Humans ; Protein Binding ; drug effects ; Protein Interaction Maps ; drug effects ; Proteins ; metabolism

The BubR1 mitotic-checkpoint protein monitors proper attachment of microtubules to kinetochores, and links regulation of chromosome-spindle attachment to mitotic-checkpoint signaling. Thus, disruption of BubR1 activity results in a loss of checkpoint control, chromosomal instability caused by a premature anaphase, and/or the early onset of tumorigenesis. The mechanisms by which deregulation and/or abnormalities of BubR1 expression operate, however, remain to be elucidated. In this study, we demonstrate that levels of BubR1 expression are significantly increased by demethylation. Bisulfite sequencing analysis revealed that the methylation status of two CpG sites in the essential BubR1 promoter appear to be associated with BubR1 expression levels. Associations of MBD2 and HDAC1 with the BubR1 promoter were significantly relieved by addition of 5-aza-2'-deoxycytidine, an irreversible DNA methyltransferase inhibitor. However, genomic DNA isolated from 31 patients with colorectal carcinomas exhibited a +84A/G polymorphic change in approximately 60% of patients, but this polymorphism had no effect on promoter activity. Our findings indicate that differential regulation of BubR1 expression is associated with changes in BubR1 promoter hypermethylation patterns, but not with promoter polymorphisms, thus providing a novel insight into the molecular regulation of BubR1 expression in human cancer cells.
Azacitidine/pharmacology ; Base Sequence ; Cell Line, Tumor ; *DNA Methylation/drug effects ; DNA Mutational Analysis ; DNA-Binding Proteins/metabolism ; *Gene Expression Regulation, Neoplastic/drug effects ; Hela Cells ; Histone Deacetylases/metabolism ; Humans ; Jurkat Cells ; Molecular Sequence Data ; Neoplasms/*genetics/*pathology ; Polymorphism, Genetic/drug effects ; Promoter Regions, Genetic/drug effects/*genetics ; Protein Binding/drug effects ; Protein Kinases/*genetics ; Protein-Serine-Threonine Kinases ; Transcription, Genetic/drug effects

OBJECTIVEThe theory of treating heart and brain simultaneously is from the theory of traditional Chinese medicine, and there aren't enough explanations for this theory from the perspective of molecular mechanism. As one successful case of this theory, the Chinese medicine formula--Buchang Naoxintong can achieve the goal of treating coronary heart disease and stroke at the same time. To illustrate the mechanism of the theory of treating heart and brain simultaneously, it is necessary to find out the molecular mechanism of this formula.

METHODUsing the network analysis method, together with two data mining methods-clustering and apriori algorithm, the frequent gene combinations interfered by the chemicals of the formula based on the protein-protein interaction networks related with coronary heart disease and stroke disease were figured out respectively. To find out the molecular mechanism of the theory of treating heart and brain simultaneously, the results got from two diseases were compared and analyzed.

RESULTBased on comparing two results from these two different diseases, the mechanism of the theory of treating heart and brain simultaneously was explained from molecular level by finding out key genes targeted by the components of this formula for both diseases and some particular genes interfered by the components for each disease. In addition, genes interfered indirectly by the chemicals for different diseases were found out based on the protein-protein interaction network.

CONCLUSIONIt can help to explain the molecular mechanism of the theory by our methods. By finding out the molecular mechanism of this theory, it can promote the progress of combination of Chinese traditional and Western medicine.

Coronary Disease ; drug therapy ; genetics ; metabolism ; Data Mining ; Drugs, Chinese Herbal ; administration & dosage ; Humans ; Protein Binding ; Protein Interaction Maps ; drug effects ; Stroke ; drug therapy ; genetics ; metabolism