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Results for “Protein expression”

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PubMed2002

Implication of protein kinase C in the regulation of DNA mismatch repair protein expression and function.

The DNA mismatch repair (MMR) proteins are essential for the maintenance of genomic stability of human cells. Compared with hereditary or even sporadic carcinomas, MMR gene mutations are very uncommon in leukemia. However, genetic instability, attested by either loss of heterozygosity or microsatellite instability, has been extensively documented in chronic or acute malignant myeloid disorders. This observation suggests that in leukemia some internal or external signals may interfere with MMR protein expression and/or function. We investigated the effects of protein kinase C (PKC) stimulation by 12-O-tetradecanoylphorbol-13-acetate (TPA) on MMR protein expression and activity in human myeloid leukemia cell lines. First, we show here that unstimulated U937 cells displayed low level of PKC activity as well as MMR protein expression and activity compared with a panel of myeloid cell lines. Second, treatment of U937 cells with TPA significantly increased (3-5-fold) hMSH2 expression and, to a lesser extent, hMSH6 and hPMS2 expression, correlated to a restoration of MMR function. In addition, diacylglycerol, a physiological PKC agonist, induced a significant increase in hMSH2 expression, whereas chelerythrine or calphostin C, two PKC inhibitors, significantly decreased TPA-induced hMSH2 expression. Reciprocally, treatment of HEL and KG1a cells that exhibited a high level of PKC expression, with chelerythrine significantly decreased hMSH2 and hMSH6 expression. Moreover, the alteration of MMR protein expression paralleled the difference in microsatellite instability and cell sensitivity to 6-thioguanine. Our results suggest that PKC could play a role in regulating MMR protein expression and function in some myeloid leukemia cells.

Base Pair Mismatch↗

PubMed2000

Adherence of Porphyromonas gingivalis to gingival epithelial cells: modulation of bacterial protein expression.

The protein profiles of Porphyromonas gingivalis (ATCC 33277 and W83) bound to KB gingival epithelial cells were analyzed by SDS-PAGE and immunoblotting. We found that a 51-kDa component was formed in bacteria that adhered to the KB cells, whereas 26- to 29-kDa bands were less intensive, in contrast to the protein profile of free bacteria. P. gingivalis ATCC 33277 incubated with protease-treated KB cells retained the profile of free bacteria. These results demonstrate the specificity of bacterial recognition of eukaryotic membrane components.

Adhesins, Bacterial↗

PubMed2006

Chlamydomonas reinhardtii: a protein expression system for pharmaceutical and biotechnological proteins.

Recombinant proteins have become more and more important for the pharmaceutical and chemical industry. Although various systems for protein expression have been developed, there is an increasing demand for inexpensive methods of large-scale production. Eukaryotic algae could serve as a novel option for the manufacturing of recombinant proteins, as they can be cultivated in a cheap and easy manner and grown to high cell densities. Being a model organism, the unicellular green alga Chlamydomonas reinhardtii has been studied intensively over the last decades and offers now a complete toolset for genetic manipulation. Recently, the successful expression of several proteins with pharmaceutical relevance has been reported from the nuclear and the chloroplastic genome of this alga, demonstrating its ability for biotechnological applications.

Animals↗

PubMed1999

[Effects of hexamethylene bisacetamide on P53 protein expression of MEC-1 cell line and cell differentiation].

OBJECTIVE: To investigate the relationship between the differentiation of the mucoepidermoid carcinoma cell line MEC-1 induced by HMBA and the expression of P53 protein. METHODS: Effects of HMBA on the growth of MEC-1 cells in vitro were measured with MTT colorimetric assay, and the content of DNA in cells was assayed by using Feulgen's staining. The P53 protein expression of MEC-1 cells was detected with ABC immunohistochemistry with anti-mutant P53 protein antibody, and the distribution of cells in different differentiation phase was determined by flow cytometry. RESULTS: HMBA inhibited the growth of MEC-1 cells in a dose-dependent and time-dependent manner, and HMBA-treated MEC-1 cells were arrested in the G1 phase and had a lower content of DNA and a lower level of P53 protein compared with the control group. CONCLUSION: The differentiation of MEC-1 cells induced by HMBA may be associated with the regulation of HMBA on the P53 protein expression and the cell differentiation cycle.

Acetamides↗

PubMed2006

[Effect of zinc deficiency on the protein expression of vitamin D receptor and calcium binding protein in growth-stage rats duodenal mucosa].

OBJECTIVE: To study the effect of zinc deficiency on the protein expression of vitamin D receptor (VDR) and calcium binding protein (CaBP) in growth-term rats duodenal mucosa and to clarify the mechanism of zinc deficiency affecting the calcium absorption by gene transcription. METHODS: Thirty weaning male rats were randomly divided into three groups: zinc deficiency (ZD), paired-fed (PF) and zinc adequation (ZA). The ZA group received a diet containing 29.5 microg/g diet ad libitum; the ZD group received a diet containing less than 0.4 microg zinc/g diet ad libitum. To eliminate the effect of zinc deficiency on appetite, the PF group received a zinc-adequate diet restricted to the quantity of food consumed the previous day by the ZD rats. After 15 days feeding, duodenal mucosa was taken to measure expression of the protein of VDR and of CaBP by immunohistochemistry and Western-blotting. RESULTS: Immunohistochemistry demonstrated that the intestinal mucosal expression of both VDR protein and CaBP protein in ZD rats significantly decreased. Analysis of the photographs showed that the number of cells expressed VDR protein in ZD rats was significantly less than that of the PF and ZA rats (P < 0.001). The number of cells which expressed VDR protein in ZD, PF, and ZA groups was 52, 162, and 220, and the number of cells which expressed CaBP protein was 169, 240 and 280 (F = 132 and 22, P < 0.001). Western-blotting showed similar results. CONCLUSIONS: Zinc deficiency, by changing the activity of VDR, changes the protein expression of VDR, and thus affects the transcription of the target gene CaBP, resulting in the absorption of calcium that causes allo-osteogenesis.

Animal Nutritional Physiological Phenomena↗

PubMed2004

[Relevant factors of nm23 protein expression and dangerous factors of bone metastasis in breast cancer].

OBJECTIVE: To investigate the relevant factors of the expression of nm23 protein and the dangerous factors of bone metastasis in breast cancer. METHODS: Seventy-six breast cancer patients confirmed by histological examination after surgeries were enrolled in this study. nm23 protein expressions in original breast cancer tissues were detected by immunohistochemical procedures. The relevant factors of nm23 protein expression and the dangerous factors of bone metastasis were conducted logistic regression analysis. RESULTS: Among the 58 breast cancer patients who did not have bone metastasis in the observation period, 55 did not have bone metastasis;while the other 18 breast cancer patients having bone metastasis were confirmed in only 14 patients. The correction was 94.83% and 77.78% respectively. The general correction was 90.79%. CONCLUSION: The detection of nm23 protein is helpful to evaluate prognosis and improve the therapy. It is one of the important methods to instruct the breast cancer patients to perform radio-nuclide imaging in the follow-up.

Adult↗

PubMed2006

Mobile phone radiation causes changes in gene and protein expression in human endothelial cell lines and the response seems to be genome- and proteome-dependent.

We have examined in vitro cell response to mobile phone radiation (900 MHz GSM signal) using two variants of human endothelial cell line: EA.hy926 and EA.hy926v1. Gene expression changes were examined in three experiments using cDNA Expression Arrays and protein expression changes were examined in ten experiments using 2-DE and PDQuest software. Obtained results show that gene and protein expression were altered, in both examined cell lines, in response to one hour mobile phone radiation exposure at an average specific absorption rate of 2.8 W/kg. However, the same genes and proteins were differently affected by the exposure in each of the cell lines. This suggests that the cell response to mobile phone radiation might be genome- and proteome-dependent. Therefore, it is likely that different types of cells and from different species might respond differently to mobile phone radiation or might have different sensitivity to this weak stimulus. Our findings might also explain, at least in part, the origin of discrepancies in replication studies between different laboratories.

Cell Line↗

PubMed2006

Recurrent NMYC copy number gain and high protein expression in basal cell carcinoma.

Formation of basal cell carcinoma (BCC) has been linked to deregulation in the sonic hedgehogh (Shh) signalling pathway. Though mutations of the genes, PTCH1 and SMO, are known to be involved in aberrant Shh signalling, the distinct downstream effectors of these genes are poorly described. Studies have indicated that the NMYC oncogene is a potential Shh downstream effector. To assess the expression of Nmyc protein and gene copy numbers of the NMYC gene locus in a representative BCC tumour collection, immunohistochemistry (IHC) and fluorescence in situ hybridisation (FISH) were performed on 273 BCC specimens of different growth patterns and anatomic localisations on tissue microarray (TMA) sections. High Nmyc protein expression was detected in 72.7% (160/220) of all BCC specimens. Strong Nmyc immunopositivity was more frequently found in infiltrative BCCs compared to nodular/superficial BCCs (p=0.005), and in BCCs of the head compared to BCCs of other anatomic localisations (p=0.021). The prevalence of NMYC copy number gains was 17.5% (37/211), including three tumours with nodular differentiation that exhibited a distinct high-level amplification of the NMYC locus. These data indicate that high expression of the Shh downstream mediator, Nmyc, is a frequent event in BCC, predominantly in more aggressive subtypes. Although the NMYC copy number gain found in a subset of cases might contribute to this aberrant Nmyc protein expression by a gene dosage effect, our data suggests that Nmyc protein can also be induced by aberrant Shh signalling, acting as an effector molecule of the Shh pathway. Novel systemic anti-sense NMYC inhibition strategies could be a promising option for therapy-refractory BCC.

Basal Cell Carcinoma↗

PubMed2001

Identification of developmentally expressed proteins that functionally interact with Nedd4 ubiquitin ligase.

Nedd4 is a HECT domain-containing ubiquitin ligase that mediates ubiquitylation and proteasome degradation of target proteins. The molecular basis for the interaction of Nedd4 with substrates lies in its WW domains, which can bind proline-rich (PY) domains in target proteins. Nedd4 is a developmentally expressed protein and may have a fundamental role to play in embryonic processes. However, whether Nedd4 has such a function is currently unknown, in part because few developmentally regulated ubiquitylation substrates have been identified or characterized. We have carried out a yeast two-hybrid screen and identified four proteins expressed in the mid-gestation embryo that are able to interact with Nedd4. Characterization of their functional interaction with Nedd4 in vitro and in vivo demonstrated that three of the four are bona fide Nedd4 binding partners, and two have the capacity to be ubiquitylation substrates. One of these is the first identified nonviral substrate for Nedd4-mediated monoubiquitylation. Interestingly, neither of these two ubiquitylated proteins interacts with Nedd4 through PY-mediated mechanisms. For one of the three Nedd4 binding partners, there was no discernable evidence of ubiquitylation. However, this protein clearly associates with Nedd4 through its PY domains and can alter the location of Nedd4 in cells, suggesting a role other than as a ubiquitylation substrate.

Blotting, Western↗

PubMed2002

Regulation of Id1 protein expression in mouse embryo fibroblasts by the type 1 insulin-like growth factor receptor.

The activated type 1 insulin-like growth factor (IGF-IR) increases the expression of Id1 proteins in mouse embryo fibroblasts (MEF). Up-regulation depends on a functional receptor and on multiple pathways originating from different domains of the receptor. In MEF, Id1 protein expression is also up-regulated by serum and certain oncogenes. Signaling through Stat3 plays an important, but not exclusive, role in the up-regulation of Id1 protein levels. In all instances, the increase in Id1 protein expression is paralleled by a corresponding increase in Id1 promoter activity, as measured with a reporter gene.

Animals↗

PubMed2006

Impaired gene and protein expression of exocytotic soluble N-ethylmaleimide attachment protein receptor complex proteins in pancreatic islets of type 2 diabetic patients.

Exocytosis of insulin is dependent on the soluble N-ethylmaleimide attachment protein receptor (SNARE) complex proteins in the B-cells. We assessed insulin release as well as gene and protein expression of SNARE complex protein in isolated pancreatic islets of type 2 diabetic patients (n = 4) and nondiabetic control subjects (n = 4). In islets from the diabetic patients, insulin responses to 8.3 and 16.7 mmol/l glucose were markedly reduced compared with control islets (4.7 +/- 0.3 and 8.4 +/- 1.8 vs. 17.5 +/- 0.1 and 24.3 +/- 1.2 microU . islet(-1) . h(-1), respectively; P < 0.001). Western blot analysis revealed decreased amounts of islet SNARE complex and SNARE-modulating proteins in diabetes: syntaxin-1A (21 +/- 5% of control levels), SNAP-25 (12 +/- 4%), VAMP-2 (7 +/- 4%), nSec1 (Munc 18; 34 +/- 13%), Munc 13-1 (27 +/- 4%), and synaptophysin (64 +/- 7%). Microarray gene chip analysis, confirmed by quantitative PCR, showed that gene expression was decreased in diabetes islets: syntaxin-1A (27 +/- 2% of control levels), SNAP-25 (31 +/- 7%), VAMP-2 (18 +/- 3%), nSec1 (27 +/- 5%), synaptotagmin V (24 +/- 2%), and synaptophysin (12 +/- 2%). In conclusion, these data support the view that decreased islet RNA and protein expression of SNARE and SNARE-modulating proteins plays a role in impaired insulin secretion in type 2 diabetic patients. It remains unclear, however, to which extent this defect is primary or secondary to, e.g., glucotoxicity.

Aged↗

PubMed2000

[MDM2 gene expression in nasopharyngeal carcinoma and its relationship with p53 protein expression and EB virus latent infection].

OBJECTIVE: To define the expression levels of MDM2 gene in nasopharyngeal carcinoma (NPC) and its relationship with p53 protein expression and EB virus latent infection. METHOD: MDM2 gene expression at mRNA and protein levels, p53 protein and EB virus DNA were detected by nonradioactive in situ hybridization (ISH), immunohistochemistry(IHC) and polymerase chain reaction (PCR) separately in 46 cases of NPC tissues and 12 cases of chronic inflammation of nasopharyngeal epithelium (CINE). RESULT: Fourteen cases of NPC showed MDM2 mRNA and protein overexpression, 38 cases were p53 protein positive, and 43 cases were EBV-DNA positive. Neither MDM2 nor p53 protein was expressed in any case of CINE. MDM2 expression was significantly related to p53 protein expression (P < 0.05), but not to EB virus latent infection in NPC. CONCLUSION: MDM2 gene may play an important role in the pathogenesis of NPC through interacting with p53 protein.

DNA, Viral↗

PubMed1997

DNA content and p53 protein expression in ductal breast cancer.

The DNA content of 85 ductal breast cancers of different histological grades was evaluated using static cytometry and correlated with immunocytochemical expression of p53 protein in tumour cells in cytological material. A statistically significant difference was observed between p53 protein expression and grade of malignancy (P < 0.001). The percentage of euploid tumours significantly decreased from grade I through grade II to grade III tumours (P < 0.001). Clonal DNA heterogeneity was observed in 26.6% of cases analysed and was correlated with p53 protein expression (P < 0.001). These changes probably reflect genomic alterations which may affect potential malignancy of breast cancer.

Adult↗

PubMed1997

Significance of basic fibroblast growth factor and fibroblast growth factor receptor protein expression in the formation of fibrotic focus in invasive ductal carcinoma of the breast.

A fibrotic focus (FF) is a scar-like area within invasive ductal carcinoma (IDC) of the breast, and has been shown to be a marker of high aggressiveness of IDC. In order to investigate the mechanism of FF formation in IDC, expression of basic fibroblast growth factor (bFGF) and fibroblast growth factor receptor (FGFR) was studied. One hundred and forty-nine IDCs were divided into solid tumors and scirrhous tumors. Immunohistochemistry was used to determine the expression of bFGF and FGFR proteins in both tumor cells and fibroblasts forming FF. Scirrhous tumors with FF showed a significantly higher frequency of bFGF protein expression than those without (P = 0.017), whereas, in solid tumors, the presence of FF was not significantly associated with the frequency of bFGF protein expression (P = 0.143). In addition, scirrhous tumors showed a significantly higher frequency of FGFR protein expression than solid tumors (P = 0.001). Among IDCs having FF and expressing bFGF protein, a significantly larger number of fibroblasts expressing FGFR protein within FF was observed in scirrhous tumors than in solid tumors (P = 0.016). The results of this study suggest that in scirrhous tumors the interaction between tumor cells and stromal fibroblasts plays an important role in the formation of FF, and that there is a paracrine mechanism between bFGF protein from tumor cells and FGFR protein in fibroblasts.

Adenocarcinoma, Scirrhous↗

PubMed2005

Enhancing protein expression in single HEK 293 cells.

Recombinant proteins are routinely expressed in heterologous expression systems such as human embryonic kidney 293 (HEK 293) cells. The efficiency of the expression is critical when the expressed protein must be characterized at the single-cell level. Here we describe a simple method by which the protein expression efficiency in single HEK 293 cells is enhanced by coexpressing simian virus 40 large T antigen (TAg), a powerful oncoprotein. Using the GluR2 ionotropic glutamate receptor as an example, we found that the receptor expression in single HEK 293S cells increased approximately seven-fold. The ratio of the plasmid amount of TAg to that of the receptor was optimized at 1:10, while the receptor function was unaffected in the presence of TAg. We further used fluorescence imaging from a population of cells as an independent detection method and found a similar increase in expression of green fluorescent protein (GFP) by TAg coexpression. This method is thus applicable for enhancing the expression of both membrane and soluble proteins at the single-cell level. More importantly, the function of a protein can be studied directly in intact cells, a feature particularly useful for studying membrane proteins.

Antigens, Viral, Tumor↗

PubMed2006

Focal S100A4 protein expression is an independent predictor of development of metastatic disease in cystectomized bladder cancer patients.

OBJECTIVE: The prognosis of patients with apparently localized, operable, muscle-invasive bladder cancer depends to a large extent on the presence or absence of subclinical, microscopic distant metastases at the time of surgery. Expression of the S100A4 protein has been shown to correlate with the risk of metastasis in both animal tumour-model systems and clinical investigations in other tumour types. The purpose of the present study was to investigate the prognostic potential of S100A4 protein expression for predicting distant metastatic relapse in muscle-invasive bladder cancer. METHODS: We analyzed 108 consecutive patients, treated for transitional cell bladder cancer with preoperative radiotherapy and cystectomy. Pretherapeutic biopsies of the bladder tumours were investigated for immunohistochemical expression of S100A4 protein and results, along with clinical and histopathological data, compared with the pattern of relapses over a 10+ yr follow-up period. RESULTS: Focal S100A4 protein expression emerged as the only significant independent predictor of distant metastatic relapse and distant metastasis-free survival in multivariate analysis. CONCLUSION: There is a potential role for this marker in denoting patients with high or low risk of distant relapse independent of clinical stage and grade.

Carcinoma, Transitional Cell↗

PubMedDate not supplied

Cytotoxic protein expression in non-Hodgkin's lymphomas and Hodgkin's disease.

Recent studies have shown that some peripheral T-cell lymphomas (PTCL) could be derived from lymphocytes with cytotoxic potential. Therefore, we have investigated by immunohistochemistry 34 cases of PTCL including 2 cases of hepatosplenic gamma delta PTCL and 5 cases of sinonasal NK-cell lymphomas as well as 7 cases of T-lymphoblastic lymphomas (T-LBL) for the expression of the cytotoxic proteins TIA-1 and granzyme B. In addition, 50 cases of Hodgkin's disease (HD) were investigated in order to see if these cytotoxic proteins are expressed by Hodgkin and Reed-Sternberg (HRS) cells. Expression of the TIA-1 is characteristic of cytotoxic cells regardless of their activation status whereas expression of granzymes is highly increased in activated cytotoxic cells. All the five cases of sinonasal NK-cell lymphomas expressed TIA-1 and granzyme B in most tumour cells. The two gamma delta PTCL cases expressed TIA-1 protein in most tumour cells but not granzyme B. Of the 32 other PTCL, 9 cases showed cytotoxic protein expression in tumour cells. These cases comprised 2 pleomorphic medium large cell (PML) (1 nodal and 1 intestinal) and 7 CD30 positive anaplastic large cell lymphomas (ALCL) (5 nodal and 2 cutaneous). Cytotoxic protein expression in our series appeared to be related to the location since 10/18 (55%) extranodal PTCL and NK-NHL and only 6/21 (28%) nodal PTCL expressed TIA-1, and related to histology since, in nodal PTCL, this pattern was observed in most anaplastic (5/6 cases) and in a few pleomorphic (1/9 cases) lymphomas, but not in AILD-type NHL (0/6 cases). The 7 cases of T-LBL did not express cytotoxic proteins in tumour cells. EBV was detected by EBER RNA in situ hybridization (RISH) in tumour cells in all 5 sinonasal NK-NHL and in scattered atypical cells in all 6 cases of AILD. Two of the 50 cases of HD weakly expressed TIA-1 and granzyme B in a proportion of HRS cells. EBV was detected by RISH in 19/50 cases of HD but no correlation was found between EBV status and expression of cytotoxic proteins in HRS cells. However, the finding that granzyme B positive cells were found very rarely in close vicinity of HRS cells suggests that the function of activated cytotoxic cells is locally inhibited by the HRS cells and/or the reactive cells in the vicinity of HRS cells. Taken together our data suggest that: a) sinonasal NK-cell NHL represent tumours of activated cytotoxic NK-cells, b) the hepatosplenic gamma delta PTCL represent tumours of nonactivated cytotoxic gamma delta T-cells, c) a small proportion of other PTCL, mostly anaplastic large cell lymphomas represent tumours of cytotoxic T-cells and d) only very few cases of HD expressing cytotoxic proteins in a proportion of tumour cells, could be derived from activated cytotoxic cells.

Granzymes↗

PubMed2004

Protein-trap version 2.1: screening for expressed proteins in mammalian cells based on their localizations.

BACKGROUND: "Protein-trap" is a method that allows epitope-tagging of endogenous proteins. This method allows for the identification of endogenously expressed proteins that exhibit specific localization of interest. This method has been recently reported for its application in the study of Drosophila development by using a relatively large epitope, green-fluorescent-protein (GFP). RESULT: Herein, we report a new "protein-trap" vector for mammalian cells. This new method utilizes a much smaller epitope-tag and also allows for drug-selection prior to the epitope-tagging. Pre-selection by drug resulted in the highly efficient protein-trapping frequency. CONCLUSION: The "protein-trap" method based on this new vector is expected to serve as a complimentary approach to the previously reported GFP-based method.

Antibodies, Monoclonal↗