Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.14.14.3 (luciferase)
 38,195 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Epidermal growth factor (EGF), determined by immunoprecipitation and Western blot analysis, increased both enzyme activity and protein level of 12-lipoxygenase in the solubilized microsomes of human epidermoid carcinoma A431 cells, respectively. The EGF-induced expression of 12-lipoxygenase mRNA was inhibited by transcription inhibitors such as actinomycin D and 5,6-dichlorobenzimidazole riboside. Promoters of different lengths for human 12-lipoxygenase gene were used to prepare the luciferase fusion vectors. These construct plasmids were transiently transfected into A431 cells, and the induction of luciferase expression by EGF was examined. A 4- to 6-fold increase in luciferase reporter activity stimulated by EGF for 18 h treatment was observed in plasmids with the 5'-flanking region length of -951 bp and that of -224 bp upstream from translation starting site. The time-dependent induction of luciferase activity by EGF paralleled the EGF-induced enzyme activity and expression of 12-lipoxygenase protein. Taken together, the results of this study indicate that EGF enhanced the transcription of the human 12-lipoxygenase gene, resulting in an increase in the amount and activity of 12-lipoxygenase.
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PMID:Epidermal growth factor enhances transcription of human arachidonate 12-lipoxygenase in A431 cells. 902 53

The effect of transient transfection with expression vectors of Ha-ras on the promoter activity of 12-lipoxygenase in human epidermoid carcinoma A431 cells was studied. Overexpression of Ha-ras increased the promoter activity in a dose- and time-dependent manner, which correlated closely with the cellular expression of Ras protein. Promoters of different gene lengths for human 12-lipoxygenase were used to prepare the luciferase fusion vectors. Following transfection by Ha-ras for 68 h, an approx. 40-fold increase in luciferase reporter activity was observed in plasmids with the 5'-flanking region ranging from -951 to -224 bp upstream from translation starting site. There was no obvious stimulation in cells transfected with a vector-bearing promoter with a length of -100 bp. These results indicate that the promoter region ranging from -224 to -100 bp was important for the Ha-ras response. With the aid of additional 5'-deletion and site-directed mutagenesis, three Sp1 binding sequences residing at -169 to -161 bp, -158 to -150 bp and -123 to -114 bp were found to be critical for the Ha-ras response of activating the transcription of human 12-lipoxygenase gene.
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PMID:Overexpression of Ha-ras enhances the transcription of human arachidonate 12-lipoxygenase promoter in A431 cells. 905 17

UCN-01 (7-hydroxyl-staurosporine) was originally isolated as a Ca2+- and phospholipid-dependent protein kinase C selective inhibitor and now is being developed as an anticancer agent. Results from our and other laboratories have suggested that UCN-01 induces preferential G1-phase accumulation in several human tumor cell lines tested. To elucidate this mechanism, we examined the effects of UCN-01 on several cell cycle-regulatory proteins critical for G1-S-phase transition in p53-mutated human epidermoid carcinoma A431 cells. After 24 h exposure at around 50% growth-inhibitory concentrations (IC50s), 260 and 520 nM, UCN-01 induced the accumulation of pRb (the dephosphorylated retinoblastoma protein form). The protein expression of cyclin A but not cyclin E was markedly reduced and that of cyclin D1 was partially reduced under the same condition. UCN-01 also showed the concentration-dependent inhibitions of the activity of cyclin-dependent kinase 2 (CDK2) using histone H1 and pRb as substrates in vitro (IC50, 530 and 640 nM, respectively). In addition, CDK2 activities of the cells pretreated with UCN-01 for 24 h at 260 and 520 nM were markedly inhibited, giving IC50s of far less than 260 nM. When the same cell lysates were analyzed by Western blotting for CDK2, the lower band (e.g., active and phosphorylated CDK2) was remarkably reduced, in accordance with the reduced activity. Furthermore, UCN-01 induced the expression of the CDK inhibitor p21 protein and its complex formation with CDK2 after 24 h exposure at 260 and 520 nM, whereas the expression level was very low or undetectable in untreated or DNA-damaged cells. The increase of p21 mRNA levels was also induced under the same condition. UCN-01 further increased luciferase activities in A431 cells transiently transfected with p21 promoter-luciferase reporter plasmid after 24 h exposure at 260 and 520 nM. UCN-01 also increased the expression of the CDK inhibitor p27 protein after 24 h exposure at 260 and 520 nM. These results suggest that G1-phase accumulation induced by UCN-01 is associated with dephosphorylation of Rb and CDK2 proteins as well as induction of CDK inhibitors p21 and p27.
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PMID:G1 phase accumulation induced by UCN-01 is associated with dephosphorylation of Rb and CDK2 proteins as well as induction of CDK inhibitor p21/Cip1/WAF1/Sdi1 in p53-mutated human epidermoid carcinoma A431 cells. 910 51

The functional 5' flanking region of the human 12-lipoxygenase in epidermoid carcinoma A431 cells was characterized. By a primer extension method, the transcription initiation sites were mapped at -47 adenosine, -48 guanosine and -55 guanosine upstream of the ATG translation start codon. Transient transfection with a series of 5' and 3' deletion constructs showed that the 5' flanking region spanning from -224 to -100 bp was important for the basal expression of 12-lipoxygenase gene. Gel mobility shift assays with antibodies of transcription factors showed that both Sp1 and Sp3 required highly GC-rich Sp1 sites within this region for binding. Disruption of two Sp1 recognition motifs residing at -158 to -150 bp and -123 to -114 bp by site-directed mutagenesis markedly reduced the basal 12-lipoxygenase promoter activity and abolished the retarded bands in a gel-shift assay, indicating that these two Sp1-binding sites were essential for gene expression. The same two Sp1-binding sites in this promoter region were also responsible for epidermal growth factor (EGF)-induced expression of 12-lipoxygenase gene. Moreover, EGF also induced the transcriptional activation of luciferase driven by SV40 early promoter, which contained rich Sp1-binding sites. Taken together, the results suggest that two specific Sp1 consensus sites are involved in the mediation of the basal promoter activity as well as EGF induction of the 12-lipoxygenase gene and that Sp1 and Sp3 transcription factors might have a role in their regulation.
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PMID:Transcriptional activation of human 12-lipoxygenase gene promoter is mediated through Sp1 consensus sites in A431 cells. 916 49

Phorbol 12-myristate 13-acetate (PMA) increased the expression of 12-lipoxygenase activity and mRNA in a time-dependent manner in human epidermoid carcinoma A431 cells. The increase of 12-lipoxygenase was accompanied by the increase in protein level in microsomes prepared from A431 cells. The PMA-induced expression of 12-lipoxygenase activity and mRNA was inhibited by the treatment of cells with a protein kinase C inhibitor GF 109203X. Promoters of different DNA lengths for human 12-lipoxygenase gene were used to prepare the luciferase fusion vectors. These plasmid constructs were transiently transfected into A431 cells. Following treatment of PMA for 18 h, a 4- to 5-fold increase in luciferase reporter activity was observed in plasmids with the 5'-flanking region length of -951 bp and that of -224 bp upstream from translation starting site. A time-dependent induction of luciferase activity by PMA was found to parallel the PMA-induced enzyme activity and mRNA expression. Transient transfection with a series of 5'-deletion constructs showed that the 5'-flanking region spanning from -224 to -100 bp from translation starting site played an important role for PMA response. Gel mobility shift assay and site-directed mutagenesis indicated that two Sp1 binding sequences residing at -158 to -150 bp and -123 to -114 bp were responsible for the PMA response in activating the transcription of human 12-lipoxygenase gene.
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PMID:Induction of 12-lipoxygenase expression by phorbol 12-myristate 13-acetate in human epidermoid carcinoma A431 cells. 944

The squamous cell carcinoma antigen (SCCA) has been used as a circulating tumor marker for the management of squamous cell carcinoma. SCCA consists of a small gene family of at least two in human genome (SCCA1 and SCCA2), which are tandemly arrayed on chromosome 18q21.3 and share 92% identical residues. SCCA expressions are tightly controlled in a tissue-specific manner. To investigate the role of SCCA2 in the cancer cells, we first isolated the human genomic clones, containing the promoter region of SCCA2 gene, and determined the nucleotide sequence surrounding the exon 1. The transcription start site was mapped by primer extension analysis, and a putative TATA box element was found in the 5'-flanking region. Other putative regulatory sequences, which include Ets binding sequence, NF-IL6 binding sequence and IRE consensus sequence, were also found in the region. Analysis of luciferase reporter gene expression in transient transfection showed that the promoter region of SCCA2 gene was located within the region from -424 to +47.
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PMID:Structural analysis of human SCC antigen 2 promoter. 993 63

We reported previously that tumor cells isolated from metastases of the in vitro transformed squamous cell carcinoma line Pam 212 exhibit an elevation in constitutive production of proinflmmatory cytokines interleukin (IL)-1alpha, IL-6, granulocyte-macrophage colony-stimulating factor, and KC (the murine homologue of chemokine Gro-alpha). The basis for constitutive expression of these cytokines after tumor progression in vivo is unknown. Regulation of the expression of these proinflammatory cytokines involves transcription factor nuclear factor kappaB (NF-kappaB), which can be activated by cytokines such as tumor necrosis factor (TNF)-alpha. In this study, we compared the constitutive and TNF-alpha-induced expression of proinflammatory cytokines in parental Pam 212 and metastatic LY-2 and LY-8 cell lines and determined the relationship of cytokine expression to activation of NF-kappaB. We found that the metastatic cell lines exhibited an increase in constitutive and TNF-alpha-inducible expression of proinflammatory cytokines when compared with parental Pam 212 cells. The increased cytokine expression was associated with an increase in constitutive and TNF-alpha-inducible activation of NF-kappaB as demonstrated by electrophoretic mobility shift assay and luciferase-reporter gene assay. Constitutive nuclear localization of NF-kappaB p65 was observed in LY-2 and LY-8 cells in culture and in tumor specimens but rarely in Pam 212 cells, consistent with the constitutive activation of NF-kappaB in tumor cels after selection in vivo. Induction of NF-kappaB by TNF-alpha was inhibited by the addition of protease inhibitors calpain inhibitor I and N-tosyl-phechloromethyl ketone and antioxidant 1-pyrrolidinecarbodithioic acid, whereas constitutive activation of NF-kappaB and cytokine KC mRNA expression was inhibited by N-tosyl-phechloromethyl ketone alone. Overexpression of a human Ikappa(B)alpha dominant suppresser in Pam 212 cells inhibited TNF-alpha-induced NF-kappaB binding activity and KC expression. These data indicate that activation of NF-kappaB contributes to increased expression of proinflammatory cytokines during metastatic tumor progression of squamous cell carcinoma, and that distinct mechanisms may be involved in the regulation of constitutive and TNF-alpha-induced activation of NF-kappaB in squamous cell carcinoma.
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PMID:The host environment promotes the constitutive activation of nuclear factor-kappaB and proinflammatory cytokine expression during metastatic tumor progression of murine squamous cell carcinoma. 1041 16

Transforming growth factor-alpha (TGF-alpha) increased the expression of 12-lipoxygenase activity in a time-dependent manner in human epidermoid carcinoma A431 cells. The increase of 12-lipoxygenase activity was accompanied by an increase in 12-lipoxygenase mRNA. The effect of TGF-alpha on the promoter activation of 12-lipoxygenase gene was analyzed by using the luciferase fusion vectors. A dose-dependent effect of TGF-alpha on the reporter activity was observed, which paralleled with its effect on enzyme activity. Transient transfection with a series of 5'-deleted constructs showed that the 5'-flanking region spanning from -224 to -100 bp from translation starting site played an important role for TGF-alpha response. Site-directed mutagenesis and gel mobility shift assay indicated that two Sp1 binding sequences residing at -158 to -150 bp and 123 to -114 bp were responsible for the TGF-alpha in activation of human 12-lipoxygenase gene transcription. Expression of Sp1, but not Sp3, stimulated the promoter activity of 12-lipoxygenase in SL2 cells, indicating that the binding of Sp1 with Sp1 binding sequences played a significant role in the regulation of 12-lipoxygenase gene.
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PMID:Induction of 12-lipoxygenase expression by transforming growth factor-alpha in human epidermoid carcinoma A431 cells. 1042 82

Promoter activation in the expression of phospholipid hydroperoxide glutathione peroxidase (PHGPx) gene in human epidermoid carcinoma A431 cells was studied in the present investigation. Luciferase reporter assays with plasmids carrying a 400 bp of the promoter DNA were performed to analyze the regulatory element in the proximal promoter of human PHGPx gene. Transient transfection with a series of 5'-deletion and internal truncation mutants showed that the 5'-flanking region spanning from -212 to -121 bp was important for the basal expression of PHGPx gene in A431 cells. A region from -170 to -140 bp was protected in DNase I footprinting assays and bound the nuclear proteins in electrophoretic mobility shift assays. This region, denoted FP3, contains the consensus recognition sites for AP-2, CCAAT-box and CRE. The oligonucleotide competitor with the mutation at CCAAT-box could not eliminate the nuclear protein binding in gel-shift assay and the site-directed mutagenesis at the CCAAT-box decreased the luciferase activity of PHGPx promoter for approximate 50% in reporter gene assays. Competition experiments indicate that the binding of nuclear factor to the FP3 region was abolished by oligodeoxyribonucleotide corresponding to NF-Y/CP1 binding site to a greater extent than by those corresponding to sites for CTF/NFI and C/EBP. Taken together, the CCAAT-box in the promoter ranging from -156 to -151 bp, bound to NF-Y/CP1, was essential for the basal expression of human PHGPx gene in A431 cells.
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PMID:The CCAAT-box binding factor NF-Y is required for the expression of phospholipid hydroperoxide glutathione peroxidase in human epidermoid carcinoma A431 cells. 1042 83

Codon 273 is one of the hot spots of missense mutation of the p53 tumor suppressor gene found in human cancers. We have previously reported that a mutation at codon 273, p53-273L (Arg --> Leu), suppresses cell growth despite its having no p53-specific transactivation activity. To further elucidate the mechanism of growth suppression caused by p53-273L, we used squamous cell carcinoma cell line HSC3 to isolate subclones containing Zn2+-inducible wild-type (wt) p53, p53-175H, and p53-273L. Northern blot hybridization of the HSC3 cells possessing an inducible function of p53 as well as a luciferase assay for the p21Waf1/Cip1/Sdi1 promoter showed that only wt p53 could induce p21Waf1/Cip1/Sdi1 transcription. Meanwhile, the expression of bax remained unchanged between, before, and after the induction of any analyzed p53s. When wt p53 was induced in HSC3 cells cultured in medium containing 5% fetal bovine serum, cell growth was suppressed through G1 arrest. On the other hand, in medium with 0.1% fetal bovine serum, the growth of HSC3 cells expressing p53-273L was suppressed to a greater degree than that of cells expressing wt p53. Flow cytometric analysis and DNA ladder formation revealed that, unlike wt p53-SN3- and p53-175H-expressing HSC3 cells, p53-273L-expressing cells contained a larger sub-G1 fraction under this culture condition. These findings suggest that p53-273L can induce apoptosis in HSC3 cells without transactivation of p21Waf1/Cip1/Sdi1 and bax.
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PMID:Induction of apoptosis by the p53-273L (Arg --> Leu) mutant in HSC3 cells without transactivation of p21Waf1/Cip1/Sdi1 and bax. 1048 21


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