Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.10.2 (focal adhesion kinase)
 44,029 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

This study investigates the feasibility of using the enhanced cyan mutant of green fluorescent protein (ECFP) as a probe for two-photon fluorescence correlation spectroscopy (FCS). Molecular dynamics and other properties of ECFP and an ECFP-tubulin fusion protein were investigated in living Potorous tridactylis (PTK2) cells. ECFP has high molecular brightness in the nucleus (eta=3.3 kcpsm) and in the cytoplasm (3.2 kcpsm) under our experimental conditions. The diffusion constants of ECFP were determined to be 20+/-7 microm(2)/s in the nucleus and 21+/-8 microm(2)/s in the cytoplasm. ECFP has stable molecular characteristics with negligible photobleaching and photodynamic effects in our measurements. At the highest concentration of monomer ECFP (425 nM) the amount of dimer ECFP was estimated to be negligible ( approximately 1.8 nM), consistent with our data analysis using a single species model. ECFP-tubulin has a diffusion constant of 6 microm(2)/s in the living cells. In addition, we demonstrate that analysis of the molecular brightness can provide a new avenue for studying the polymerization state of tubulin. We suggest that the tubulin in the vicinity of the nucleus exists primarily as a heterodimer subunit while those in the area away from the nucleus (d>5 microm) are mostly oligomers. We conclude that ECFP is a useful genetic fluorescent probe for FCS studies of various cellular processes when in fusion to other biomolecules of interest.
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PMID:Fluorescence correlation spectroscopy investigation of a GFP mutant-enhanced cyan fluorescent protein and its tubulin fusion in living cells with two-photon excitation. 1506 7

Mitochondrial dysfunction is a hallmark of cancer cells. However, genetic response to mitochondrial dysfunction during carcinogenesis is unknown. To elucidate genetic response to mitochondrial dysfunction we used Saccharomyces cerevisiae as a model system. We analyzed genome-wide expression of nuclear genes involved in signal transduction and transcriptional regulation in a wild-type yeast and a yeast strain lacking the mitochondrial genome (rho(0)). Our analysis revealed that the gene encoding cAMP-dependent protein kinase subunit 3 (PKA3) was upregulated. However, the gene encoding cAMP-dependent protein kinase subunit 2 (PKA2) and the VTC1, PTK2, TFS1, CMK1, and CMK2 genes, involved in signal transduction, were downregulated. Among the known transcriptional factors, OPI1, MIG2, INO2, and ROX1 belonged to the upregulated genes, whereas MSN4, MBR1, ZMS1, ZAP1, TFC3, GAT1, ADR1, CAT8, and YAP4 including RFA1 were downregulated. RFA1 regulates DNA repair genes at the transcriptional level. RFA is also involved directly in DNA recombination, DNA replication, and DNA base excision repair. Downregulation of RFA1 in rho(0) cells is consistent with our finding that mitochondrial dysfunction leads to instability of the nuclear genome. Together, our data suggest that gene(s) involved in mitochondria-to-nucleus communication play a role in mutagenesis and may be implicated in carcinogenesis.
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PMID:Genome-wide analysis of signal transducers and regulators of mitochondrial dysfunction in Saccharomyces cerevisiae. 1512 4

To clarify the genetic aberrations involved in the development and progression of hepatitis C virus-associated hepatocellular carcinoma (HCV-HCC), we investigated DNA copy number aberrations (DCNAs) in 19 surgically resected HCCs by conventional CGH and array CGH. Conventional CGH revealed that increases of DNA copy number were frequent at 1q (79% of the cases), 8q (37%), 6p (32%), and 10p (32%) and that decreases were frequent at 17p (79%), 16q (58%), 4q (53%), 13q (42%), 10q (37%), 1p (32%), and 8p (32%). In general, genes that showed DCNAs by array CGH were usually located in chromosomal regions with DCNAs detected by conventional CGH analysis. Increases in copy numbers of the LAMC2, TGFB2, and AKT3 genes (located on 1q) and decreases in copy numbers of FGR/SRC2 and CYLD (located on 1p and 16q, respectively) were observed in more than 30% of tumors, including small, well-differentiated carcinomas. These findings suggest that these genes are associated with the development of HCV-HCC. Increases of MOS, MYC, EXT1, and PTK2 (located on 8q) were detected exclusively in moderately and poorly differentiated tumors, suggesting that these alterations contribute to tumor progression. In conclusion, chromosomal and array CGH technologies allow identification of genes involved in the development and progression of HCV-HCC.
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PMID:Analysis of DNA copy number aberrations in hepatitis C virus-associated hepatocellular carcinomas by conventional CGH and array CGH. 1513 72

The use of focused high-intensity light sources for ablative perturbation has been an important technique for cell biological and developmental studies. In targeting subcellular structures many studies have to deal with the inability to target, with certainty, an organelle or large macromolecular complex. Here we demonstrate the ability to selectively target microtubule-based structures with a laser microbeam through the use of enhanced yellow fluorescent protein (EYFP) and enhanced cyan fluorescent protein (ECFP) variants of green fluorescent protein fusions of tubule. Potorous tridactylus (PTK2) cell lines were generated that stably express EYFP and ECFP tagged to the alpha-subunit of tubulin. Using microtubule fluorescence as a guide, cells were irradiated with picosecond laser pulses at discrete microtubule sites in the cytoplasm and the mitotic spindle. Correlative thin-section transmission electron micrographs of cells fixed one second after irradiation demonstrated that the nature of the ultrastructural damage appeared to be different between the EYFP and the ECFP constructs suggesting different photon interaction mechanisms. We conclude that focal disruption of single cytoplasmic and spindle microtubules can be precisely controlled by combining laser microbeam irradiation with different fluorescent fusion constructs. The possible photon interaction mechanisms are discussed in detail.
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PMID:Controlled ablation of microtubules using a picosecond laser. 1545 3

Polyamines are small charged molecules essential for various cellular functions, but at high levels they are cytotoxic. Two yeast kinases, SKY1 and PTK2, have been demonstrated to regulate polyamine tolerance. Here we report the identification and characterization of additional genes involved in regulating polyamine tolerance: YGL007W, FES1 and AGP2. Deletion of YGL007W, an open reading frame located within the promoter of the membrane proton pump PMA1, decreased Pma1p expression. Deletion of FES1 or AGP2 resulted in reduced polyamine uptake. While high-affinity spermine uptake was practically absent in agp2Delta cells, fes1Delta cells displayed only reduced affinity towards spermine. Despite the reduced uptake, the resistant strains accumulated significant levels of polyamines and displayed increased ornithine decarboxylase activity, suggesting reduced polyamine sensing. Interestingly, fes1Delta cells were highly sensitive to salt ions, suggesting different underlying mechanisms. These results indicate that mechanisms leading to polyamine tolerance are complex, and involve components other than uptake.
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PMID:Mechanism of polyamine tolerance in yeast: novel regulators and insights. 1637 85

The aim of this study was to screen genetic as well as expression alterations in prostate cancer. Array comparative genomic hybridization (aCGH) to a 16K cDNA microarray was performed to analyze DNA sequence copy number alterations in 5 prostate cancer cell lines and 13 xenografts. The aCGH confirmed the previously implicated common gains and losses, such as gains at 1q, 7, 8q, 16p and 17q and losses at 2q, 4p/q, 6q, 8p, 13q, 16q, 17p and 18q, which have previously been identified by chromosomal CGH (cCGH). Because of the higher resolution of aCGH, the minimal commonly altered regions were significantly narrowed-down. For example, the gain of 8q was mapped to three independent regions, 8q13.3-q21.11, 8q22.2 and 8q24.13-q24.3. In addition, a novel recurrent gain at 9p13-q21 was identified. The concomitant expression analysis indicated that genome-wide DNA sequence copy number (gene dosage) was significantly associated with the expression level (p < 0.0001). The analyses indicated several individual genes whose expression was associated with the gene copy number. For example, gains of PTK2 and FZD6, were associated with the increased expression, whereas losses of TNFRSF10B (alias DR5) and ITGA4 with decreased expression. In conclusion, the aCGH mapping data will aid in the identification of genes altered in prostate cancer. The combined expression and copy number analysis suggested that even a low-level copy number change may have significant effect on gene expression, and thus on the development of prostate cancer.
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PMID:Genetic aberrations in prostate cancer by microarray analysis. 1664 77

Cervical cancer (CC) cells exhibit complex karyotypic alterations, which is consistent with deregulation of numerous critical genes in its formation and progression. To characterize this karyotypic complexity at the molecular level, we used cDNA array comparative genomic hybridization (aCGH) to analyze 29 CC cases and identified a number of over represented and deleted genes. The aCGH analysis revealed at least 17 recurrent amplicons and six common regions of deletions. These regions contain several known tumor-associated genes, such as those involved in transcription, apoptosis, cytoskeletal remodeling, ion-transport, drug metabolism, and immune response. Using the fluorescence in situ hybridization (FISH) approach we demonstrated the presence of high-level amplifications at the 8q24.3, 11q22.2, and 20q13 regions in CC cell lines. To identify amplification-associated genes that correspond to focal amplicons, we examined one or more genes in each of the 17 amplicons by Affymetrix U133A expression arrays and semiquantitative reverse-transcription PCR (RT-PCR) in 31 CC tumors. This analysis exhibited frequent and robust upregulated expression in CC relative to normal cervix for genes EPHB2 (1p36), CDCA8 (1p34.3), AIM2 (1q22-23), RFC4, MUC4, and HRASLS (3q27-29), SKP2 (5p12-13), CENTD3 (5q31.3), PTK2, RECQL4 (8q24), MMP1 and MMP13 (11q22.2), AKT1 (14q32.3), ABCC3 (17q21-22), SMARCA4 (19p13.3) LIG1 (19q13.3), UBE2C (20q13.1), SMC1L1 (Xp11), KIF4A (Xq12), TMSNB (Xq22), and CSAG2 (Xq28). Thus, the gene dosage and expression profiles generated here have enabled the identification of focal amplicons characteristic for the CC genome and facilitated the validation of relevant genes in these amplicons. These data, thus, form an important step toward the identification of biologically relevant genes in CC pathogenesis. This article contains Supplementary Material available at http://www.interscience.wiley.com/jpages/1045-2257/suppmat.
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PMID:Gene dosage alterations revealed by cDNA microarray analysis in cervical cancer: identification of candidate amplified and overexpressed genes. 1724 65

Therapeutic strategies based on antiangiogenic approaches are beginning to show great promise in clinical studies. However, full realization of these approaches requires identification of key differences in gene expression between endothelial cells from tumors versus their normal counterparts. Here, we examined gene expression differences in purified endothelial cells from 10 invasive epithelial ovarian cancers and 5 normal ovaries using Affymetrix U133 Plus 2.0 microarrays. More than 400 differentially expressed genes were identified in tumor-associated endothelial cells. We selected and validated 23 genes that were overexpressed by 3.6- to 168-fold using real-time reverse transcription-PCR and/or immunohistochemistry. Among these, the polycomb group protein enhancer of Zeste homologue 2 (EZH2), the Notch ligand Jagged1, and PTK2 were elevated 3- to 4.3-fold in tumor-associated endothelial cells. Silencing these genes individually with small interfering RNA blocked endothelial cell migration and tube formation in vitro. The present study shows that tumor and normal endothelium differ at the molecular level, which may have significant implications for the development of antiangiogenic therapies.
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PMID:Gene alterations identified by expression profiling in tumor-associated endothelial cells from invasive ovarian carcinoma. 1730 18

Primary serous ovarian carcinoma (OVCA) and serous Fallopian tube carcinoma (FTC), both belonging to the BRCA-linked tumour spectrum, share many properties and are treated similarly. However, a detailed molecular comparison has been lacking. We hypothesized that comparative genomic studies of serous OVCAs and FTCs should point to gene regions critically involved in their tumorigenesis. Array comparative genomic hybridization (array CGH) analysis indicated that serous OVCAs and serous FTCs displayed common but also more distinctive patterns of recurrent changes. Targeted gene identification using a dedicated multiplex ligation-dependent probe amplification (MLPA) probe set directly identified EIF2C2 on 8q as a potentially important driver gene. Other previously unappreciated gained/amplified genes included PSMB4 on 1q, MTSS1 on 8q, TEAD4 and TSPAN9 on 12p, and BCAS4 on 20q. SPINT2 and ACTN4 on 19q were predominantly found in FTCs. Gains/amplifications of CCNE1 and MYC, often in conjunction with changes in genes of the AKT pathway, EVI1 and PTK2, seemed to be involved at earlier stages, whereas changes of ERBB2 were associated with advanced stages. The only BRCA1-mutated FTC shared common denominators with the sporadic tumours. In conclusion, the data suggest that serous OVCAs and FTCs, although related, exhibit differences in genomic profiles. In addition to known pathways, new genes/pathways are likely to be involved, with changes in an miRNA-associated gene, EIF2C2, as one important new feature. Dedicated MLPA sets constitute potentially important tools for differential diagnosis and may provide footholds for tailored therapy.
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PMID:DNA profiling of primary serous ovarian and fallopian tube carcinomas with array comparative genomic hybridization and multiplex ligation-dependent probe amplification. 1766 15

To gain further insight into alterations in cellular pathways, tumor profiling, and marker discovery in colorectal cancer (CRC) we used a new antibody microarray specific for cell signaling. Soluble protein extracts were prepared from paired tumor/normal biopsies of 11 patients diagnosed with colorectal carcinoma at different stages; four liver carcinomas were used as a reference. Antibody microarray analysis identified 46 proteins that were differentially expressed between normal colorectal epithelium and adenocarcinoma. These proteins gave a specific signature for CRC, different from other tumors, as well as a panel of novel markers and potential targets for CRC. Twenty-four proteins were validated by using a specific colorectal cancer tissue microarray and immunoblotting analysis. Together with some previously well known deregulated proteins in CRC (beta-catenin, c-MYC, or p63), we found new potential markers preferentially expressed in CRC tumors: cytokeratin 13, calcineurin, CHK1, clathrin light chain, MAPK3, phospho-PTK2/focal adhesion kinase (Ser-910), and MDM2. CHK1 antibodies were particularly effective in discriminating between tumoral and normal mucosa in CRC. Moreover a global picture of alterations in signaling pathways in CRC was observed, including a significant up-regulation of different components of the epidermal growth factor receptor and Wnt/beta-catenin pathways and the down-regulation of p14(ARF). The experimental approach described here should be applicable to other pathologies and neoplastic processes.
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PMID:A proteomics analysis of cell signaling alterations in colorectal cancer. 1784 89


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