We reported that the RFP gene encodes a protein with putative zinc finger domains and was involved in the activation of the ret proto-oncogene. To further characterize the RFP protein, we developed a polyclonal antibody against the product synthesized from a fragment of the RFP cDNA expressed in Escherichia coli. Western blot analysis showed that RFP was identified as a 58 kDa protein in cell lysates from four human and rodent cell lines and from mouse testis. In addition, a unique 68 kDa protein was detected in the testis. Using AH7974 (rat ascites hepatoma) and Raji (human Burkitt lymphoma) cells, we demonstrated strong association of RFP with the nuclear matrix. Furthermore, RFP solubilized from the nuclear matrix had DNA-binding activity although it appears to bind more preferentially to double-stranded DNA than to single-stranded DNA. These results thus suggest that RFP may play a role in molecular processes which occur in the nuclear matrix. 相似文献
Salvelinus leucomaenis (white-spotted charr) and S. malma (Dolly Varden) are distributed throughout Hokkaido Island, Japan, but sites where they occur in sympatry are rare. In general, S. malma inhabit upstream reaches and S. leucomaenis extend downstream to the ocean. Factors influencing their distribution were analyzed at four spatial scales ranging from the whole island to individual stream pools. At the island scale, S. leucomaenis were found in the warmer south-west region and at lower altitudes elsewhere, whereas S. malma were found in the colder north-east and at higher altitudes. At a regional scale, the downstream limit of S. malma and upstream limit of S. leucomaenis shifted to lower altitude from south-west to north-east across the island, coincident with the decrease in temperature. Further analysis showed that transition points from S. leucomaenis or sympatry to S. malma in individual watersheds were closely related to an index of cumulative mean monthly temperatures exceeding 5°C. However, at the scale of a single watershed, the transition occurred at different altitudes, gradients, and temperatures in two tributaries, apparently because stream discharge, habitat, and disturbances from floods interacted with these abiotic factors to limit distribution. The two charr species developed interspecific dominance hierarchies in individual pools, and there was strong complementary density compensation among stream pools that could be explained by interspecific competition but not by differences in habitat. However, patterns at watershed and regional scales suggested that interspecific competition interacts with temperature in complex ways. We conclude that the importance of various abiotic and biotic factors in shaping Hokkaido charr distributions depends on the scale at which they are viewed. 相似文献
1. 1. The objective of this paper is to investigate the indoor environment from the viewpoint of interaction between physical environment and the human responses. The field survey has been conducted over 1 year.
2. 2. A continuous measurement has been carried out for 1 week and distribution of variables have been measured for 1 day.
3. 3. The attitude of workers was investigated by a questionnaire.
4. 4. As the result, average luminance represented more than 1000 lx in the new building, in contrast with less tha 300 lx in the existing building.
5. 5. There was a significant difference of the occupants' response to the light environment between the two buildings.
6. 6. Measured thermal conditions are on the edge of the ASHRAE comfort envelope in summer, and in the neighborhood of the lower dry limit of the envelope in spring.
7. 7. The occupants' evaluations were remarkably changed before and after the moving. The office environment is better than that of the factory.
The major obstacles to treatment of pancreatic cancer are the highly invasive capacity and resistance to chemo- and radiotherapy. Glycogen synthase kinase 3β (GSK3β) regulates multiple cellular pathways and is implicated in various diseases including cancer. Here we investigate a pathological role for GSK3β in the invasive and treatment resistant phenotype of pancreatic cancer.
Methods
Pancreatic cancer cells were examined for GSK3β expression, phosphorylation and activity using Western blotting and in vitro kinase assay. The effects of GSK3β inhibition on cancer cell survival, proliferation, invasive ability and susceptibility to gemcitabine and radiation were examined following treatment with a pharmacological inhibitor or by RNA interference. Effects of GSK3β inhibition on cancer cell xenografts were also examined.
Results
Pancreatic cancer cells showed higher expression and activity of GSK3β than non-neoplastic cells, which were associated with changes in its differential phosphorylation. Inhibition of GSK3β significantly reduced the proliferation and survival of cancer cells, sensitized them to gemcitabine and ionizing radiation, and attenuated their migration and invasion. These effects were associated with decreases in cyclin D1 expression and Rb phosphorylation. Inhibition of GSK3β also altered the subcellular localization of Rac1 and F-actin and the cellular microarchitecture, including lamellipodia. Coincident with these changes were the reduced secretion of matrix metalloproteinase-2 (MMP-2) and decreased phosphorylation of focal adhesion kinase (FAK). The effects of GSK3β inhibition on tumor invasion, susceptibility to gemcitabine, MMP-2 expression and FAK phosphorylation were observed in tumor xenografts.
Conclusion
The targeting of GSK3β represents an effective strategy to overcome the dual challenges of invasiveness and treatment resistance in pancreatic cancer. 相似文献
Mutations in the Cu/Zn-superoxide dismutase (SOD1) gene cause familial amyotrophic lateral sclerosis (ALS) through the gain of a toxic function; however, the nature of this toxic function remains largely unknown. Ubiquitylated aggregates of mutant SOD1 proteins in affected brain lesions are pathological hallmarks of the disease and are suggested to be involved in several proposed mechanisms of motor neuron death. Recent studies suggest that mutant SOD1 readily forms an incorrect disulfide bond upon mild oxidative stress in vitro, and the insoluble SOD1 aggregates in spinal cord of ALS model mice contain multimers cross-linked via intermolecular disulfide bonds. Here we show that a non-physiological intermolecular disulfide bond between cysteines at positions 6 and 111 of mutant SOD1 is important for high molecular weight aggregate formation, ubiquitylation, and neurotoxicity, all of which were dramatically reduced when the pertinent cysteines were replaced in mutant SOD1 expressed in Neuro-2a cells. Dorfin is a ubiquityl ligase that specifically binds familial ALS-linked mutant SOD1 and ubiquitylates it, thereby promoting its degradation. We found that Dorfin ubiquitylated mutant SOD1 by recognizing the Cys(6)- and Cys(111)-disulfide cross-linked form and targeted it for proteasomal degradation. 相似文献
Reactive oxygen species (ROS) are released into the alveolar space and contribute to alveolar epithelial damage in patients with acute lung injury. However, the role of ROS in alveolar repair is not known. We studied the effect of ROS in our in vitro wound healing model using either human A549 alveolar epithelial cells or primary distal lung epithelial cells. We found that H(2)O(2) inhibited alveolar epithelial repair in a concentration-dependent manner. At similar concentrations, H(2)O(2) also induced apoptosis, an effect seen particularly at the edge of the wound, leading us to hypothesize that apoptosis contributes to H(2)O(2)-induced inhibition of wound repair. To learn the role of apoptosis, we blocked caspases with the pan-caspase inhibitor N-benzyloxycarbonyl-Val-Ala-Asp (zVAD). In the presence of H(2)O(2), zVAD inhibited apoptosis, particularly at the wound edge and, most importantly, maintained alveolar epithelial wound repair. In H(2)O(2)-exposed cells, zVAD also maintained cell viability as judged by improved cell spreading and/or migration at the wound edge and by a more normal mitochondrial potential difference compared with cells not treated with zVAD. In conclusion, H(2)O(2) inhibits alveolar epithelial wound repair in large part by induction of apoptosis. Inhibition of apoptosis can maintain wound repair and cell viability in the face of ROS. Inhibiting apoptosis may be a promising new approach to improve repair of the alveolar epithelium in patients with acute lung injury. 相似文献
Amyotrophic lateral sclerosis is characterized by selective motor neuron degeneration. An apoptotic pathway is thought to be involved. It is difficult, however, to analyze the molecular pathogenic mechanism in single motor neurons because of complexity in the neural tissue, which consists of multiple lineages of cells neighboring motor neurons. We quantified the caspase-1 and -3 mRNA in single motor neurons and neighboring glial cells isolated from the spinal ventral horn of mutant SOD1 transgenic (Tg) mice and littermates. Motor neurons and neighboring glial cells were isolated from spinal sections by laser microdissection, and the mRNAs were quantified by RT-PCR. In the Tg mice, caspase-1 mRNA was first upregulated in motor neurons and second in glial cells. The caspase-3 mRNA was increased in motor neurons following the caspase-1 mRNA. These results indicated that caspase-1 and -3 mRNAs are differentially upregulated in motor neurons and glial cells of the Tg mice, and that mRNAs in isolated cells can be accurately assessed using our procedures. 相似文献