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NKX3.1是前列腺特异表达的同源盒基因,在前列腺癌的发生发展中起重要作用,而在前列腺癌进展中常会发生p53的基因突变.为研究两者之间的关系,构建NKX-3.1启动子(1 040bp)-荧光素酶报告基因重组质粒(pGL3-1040)及其缺失突变体,瞬时转染前列腺癌细胞LNCaP.通过荧光素酶表达活性分析,检测p53过表达对NKX3.1启动子活性的影响.结果表明:p53在LNCaP细胞中过表达可明显抑制NKX3.1启动子活性;RT-PCR及Western印迹检测p53过表达对NKX3.1表达的影响.结果表明,p53过表达可以明显抑制同源盒基因NKX3.1的表达.通过TRANSFAC软件分析,在NKX3.1基因上游-526至-507区存在一个p53反应元件的5′核心序列.缺失pGL3-1040中的p53反应元件核心序列并不能消除p53对NKX3.1启动子的抑制作用,表明p53不是通过p53反应元件直接抑制NKX3.1启动子活性.进一步通过5′缺失突变分析,发现NKX3.1启动子-140~+8 bp区仍受p53负调控.此148 bp区域中含有一个Sp1和一个CREB元件,瞬时共转染Sp1表达载体或CREB表达载体的结果表明,p53并不是通过与Sp1或CREB相互作用对NKX3.1启动子发挥抑制作用的.上述结果表明,p53过表达可以抑制同源盒基因NKX3.1启动子活性,下调NKX3.1基因的转录,其调控机制有待进一步研究. 相似文献
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A derivative of Azospirillum brasilense Sp245, Sp245.5, which spontaneously lost 85 and 120 MDa replicons upon the formation of a new megaplasmid, has been shown
to produce a novel lipopolysaccharide and to lose Calcofluor-binding polysaccharides. As compared to Sp245, the derivative
displays notably increased heavy metal tolerance. The phenotypes of Sp245 and Sp245.5 are characterized by the following minimal
inhibitory concentrations (MICs) of heavy metals: 0.5 and 0.9 μmol l−1 of Ag+, 0.4 and 0.7 mmol l−1 of Co2+, 0.9 and 4.7 mmol l−1 of Cu2+, and 3.1 and 11.5 mmol l−1 of Zn2+, respectively. In Sp245, in the presence of a nonlethal concentration (0.625 μmol l−1) of the efflux pump inhibitor carbonyl cyanide m-chlorophenylhydrazone (CCCP), the MIC of cobalt, copper, and zinc drop 1.3- to 1.6-fold, but the low tolerance to silver
is unaffected. In Sp245.5, CCCP does not affect cobalt tolerance, suppresses tolerance to copper and silver to the wild-type
levels, and causes a 1.4-fold decrease in resistance to zinc. Therefore, significant elevation of heavy metal tolerance in
Sp245.5 seems caused by the induction/overexpression of the proton-dependent efflux of certain metal ions. The novel cell
surface and other unknown factors could also be responsible for the increased tolerance of A. brasilense Sp245.5 to heavy metals. 相似文献
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Pengju Zhang Wenwen Liu Ju Zhang Hengyun Guan Weiwen Chen Xing Cui Qingwei Liu Anli Jiang 《Molecular biology reports》2010,37(3):1505-1512
NKX3.1, a prostate-specific gene, plays an important role in prostate development and carcinogenesis. However, its precise function
has not been established. In present study, we transfected the NKX3.1 eukaryotic expression plasmid (pcDNA3.1-NKX3.1) into
human prostate cancer cells PC-3, which lack of NKX3.1 expression, and established stable transfectants. Then, we investigated
the influence of NKX3.1 on the cell growth, cell migration and colony formation efficiency. The results showed that restoration
of NKX3.1 expression inhibited proliferation and invasion activities of PC-3 cells. Further, a cDNA microarray containing 22,000 human
genes was used to identify the gene expression differences. The results showed that there were 1,953 genes showing more than
a two-fold difference in expression. Subsequent ontological analysis revealed that a large proportion of the classified genes
were related to cell growth, cell signal and cell invasion. Finally, the expression of Caspase-3, Bcl-2, P27, Cdk6 and AMACR,
randomly selected genes from microarray data, was validated by RT-PCR and western blot. Collectively, our results first analyzed
the gene expression profile in PC-3 cells induced by NKX3.1 and indicated that NKX3.1 might exert its function by regulating
the expression of relative genes. 相似文献
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To investigate the correlation between mutations in promoter, attenuator, and the AmpC enzyme overproduction in Escherichia coli. ampC Promoters from 4 Escherichia coli clinical isolates were cloned upstream to the chloramphenicol acetyltransferase (CAT) gene in pCAT3 reporter plasmid. Promoter
strengths were measured by chloramphenicol MIC and gene sequencing was done on the cloned ampC promoter and attenuator. The strength of promoters from AmpC hyperproducers were 8- to 64-fold higher than those from a low-level
AmpC producers. In one of the high-strength promoters, the mutations were located at positions −32, +22, +26, +32 (attenuator),
−76, and +79. In another promoter, the mutations were located at positions −88, −82, −18, −1, and +58. In the third promoter,
mutations were found at positions −1, +58, −80, −73, −28, and +82. Mutations in Escherichia coli promoter and attenuator sequences promoted Chloramphenicol MICs, which may be the primary causal mechanism for resistance
to β-lactams antibiotics. 相似文献
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Zhang Pengju Wang Aiying Ni Nana Liu Qingwei Jiang Anli 《Biochemical and biophysical research communications》2010,398(3):457-214
NKX3.1, a prostate-specific homeobox gene, plays an important role in prostate cancer and usually functions as tumor suppressor gene. Previously we have demonstrated that forced expression of NKX3.1 reduced cell growth and invasion in prostate cancer cell line PC-3. Presently, we investigated the effect of NKX3.1 on the sensitivity of the prostate cancer cells to apoptosis inducer tumor necrosis factor-α (TNF-α) and cycloheximide (CHX). PC-3 cells were transfected with NKX3.1 expression plasmid (pcDNA3.1-NKX3.1) and LNCaP cells were transfected with siRNA expression plasmid (pRNAT-RNAi1) targeting NKX3.1. The cell morphology and apoptotic rate were analyzed by Hoechst 33342 staining and Flow Cytometry in absence or presence of TNF-α and CHX. The activity of caspase-3 was determined using DEVD-pNA as substrate. Simultaneously, the effect of NKX3.1 on caspase-3 expression was detected using RT-PCR and Western blot. The results showed that ectopic expression of NKX3.1 promoted TNF-α/CHX-induced apoptosis in PC-3 cells, whereas knockdown of NKX3.1 protected LNCaP cells from apoptosis induced by TNF-α/CHX. The pro-apoptosis activity of NKX3.1 might partially contribute to its elevation of caspase-3 expression and activity. Manipulating NKX3.1 expression should be a promising therapeutic strategy for treating both androgen-dependent and androgen-independent prostate cancer. 相似文献
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Faghih Z Erfani N Razmkhah M Sameni S Talei A Ghaderi A 《Molecular biology reports》2009,36(7):1923-1928
Interleukin-13 (IL-13) is a TH2 cytokine with direct and indirect immunoregulatory functions on cancer cells. The cytokine
has been reported to have some polymorphic variations at the gene level associated with some immune related diseases including
asthma and allergy. In the present study, association of three IL13 gene polymorphisms at positions −1512 A/C and −1055 C/T in the promoter and +2044 G/A in exon-4 was investigated in Iranian
women with breast cancer and healthy controls. Genotyping of IL13 gene polymorphisms were performed by PCR–RFLP methods. Serum level of IL-13 was assessed by ELISA. Haplotypes were constructed
from genotypic data using Arlequin 3.1 software package. Haplotype analysis revealed higher frequency of a three-locus haplotype,
ACA (−1512A/−1055C/+2044A), in normal women than breast cancer patients (P < 0.025). Haplotype CCA, from the other hand, was observed with more frequency among patients than controls (P < 0.03). No statistically significant differences were found in the frequency of genotypes and alleles between patients and
control group. No association was observed between investigated genotypes and other prognostic factors including tumor type,
lymph node involvement and tumor size. IL-13 serum level was undetectable in both patients and control subjects. Despite observing
no association between breast cancer and the single SNPs, results of this investigation suggest that the presence of CCA haplotype
of IL13 gene may be associated with susceptibility of Iranian women to breast cancer. 相似文献
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A recombinant gene expressing a Cry1Ac-GFP fusion protein with a molecular mass of approximately 160 kD was constructed to
investigate the expression of cry1Ac, the localization of its gene product Cry1Ac, and its role in crystal development in Bacillus thuringiensis. The cry1Ac-gfp fusion gene under the control of the cry1Ac promoter was cloned into the plasmid pHT304, and this construct was designated pHTcry1Ac-gfp. pHTcry1Ac-gfp was transformed
into the crystal-negative strain, HD-73 cry−, and the resulting strain was named HD-73−(pHTcry1Ac-gfp). The gfp gene was then inserted into the large HD-73 endogenous plasmid pHT73 and fused with the 3′ terminal of the cry1Ac gene by homologous recombination, yielding HD-73Φ(cry1Ac-gfp)3534. Laser confocal microscopy and Western blot analyses showed for the first time that the Cry1Ac-GFP fusion proteins in
both HD-73−(pHTcry1Ac-gfp) and HD-73Φ(cry1Ac-gfp)3534 were produced during asymmetric septum formation. Surprisingly, the Cry1Ac-GFP fusion protein showed polarity and was
located near the septa in both strains. There was no significant difference between Cry1Ac-GFP and Cry1Ac in their toxicity
to Plutella xylostella larvae. 相似文献
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Intercellular adhesion molecules (ICAMs) are known to be involved in various human cancers. An ICAM gene cluster lying within a 26 kb region on chromosome 19p13.2, and containing ICAM1, ICAM4, and ICAM5 has recently been identified as harboring a breast and prostate cancer susceptibility locus in two populations of European ancestry from Germany and Australia. The objective of this study was to confirm the ICAM association with prostate cancer in a sample of African American prostate cancer cases (N = 286) and controls (N = 391). Six single nucleotide polymorphisms (SNPs) within the three ICAM genes were genotyped. To control for potential population stratification an ancestry-adjusted association analysis was performed. We found that ICAM1 SNPs, −9A/C (rs5490) and K469E (rs5498) were associated with prostate cancer risk in men with a family history of prostate cancer (P = 0.008). Specifically, increased risk was observed for individuals who possessed the CC genotype of the −9 A/C variant (odds ratio = 2.5; 95% CI = 1.0–6.3) and at least one G allele of non-synonymous K469E variant (odds ratio = 1.8; 95% CI = 1.2–3.1). Strong linkage disequilibrium was observed across the ICAM region (P < 0.001). A common haplotype within the ICAM gene cluster, harboring the −9A/C variant was significantly associated with prostate cancer (P = 0.03), mainly due to men with family history (P = 0.01). Our results replicate previous findings of association of the ICAM gene cluster with prostate cancer and suggest that common genetic variation within ICAM1 and not ICAM5 may be an important risk factor for prostate cancer. 相似文献
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Ruiping Hou Bangwei Cao Zhongdong Chen Yong Li Tao Ning Chunhui Li Changqing Xu Ziping Chen 《Molecular biology reports》2010,37(1):515-520
Cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) was widely accepted as a pivotal molecule in downregulating T-cell mediated immune responses. In this study we investigated
the polymorphisms which would impact the CTLA-4 gene expression and function to assess the association with the risk of gastric cancer. 205 gastric cancer patients and 262
healthy controls were included in the case-control study. PCR and restriction fragment length polymorphism (RFLP) methods
were performed to identify the +49A/G and promoter −1661A/G polymorphisms. The promoter −1772T/C polymorphism was detected
by PCR amplification refractory mutation system (ARMS) technique. A significant difference was observed between case and control
groups. The frequency of +49A/G polymorphism AG and −1661A/G polymorphism GG genotype were significantly higher in patients
than in controls (OR = 2.15, OR = 1.88, respectively). No significant difference was found in the allelic frequency of −1772T/C
polymorphism between cases and controls (P = 0.478). By the haplotype analysis, logistic regression showed the frequency of haplotype A (GAT) and D (AGT) in the case
group revealed significant difference compared with in control group(OR = 2.00, P < 0.001; OR = 1.62, P = 0.043, respectively). Our findings implied the genetic variations within CTLA-4 gene would be a critical risk factor to the susceptibility of gastric cancer. 相似文献
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N. A. Ermolenko U. A. Boyarskikh A. G. Sushko E. N. Voronina I. A. Selezneva T. V. Sinkina A. F. Lazarev V. D. Petrova M. L. Filipenko 《Russian Journal of Genetics》2010,46(12):1486-1491
The frequencies of the polymorphic gene variants MnSOD Ala9Val, GPX1 Pro198Leu, and GSTP1 Ile105Val were estimated in female residents of Altai krai with breast cancer. The frequency distributions of the genotypes for
all genes studied in both patients and control subjects fit the Hardy-Weinberg equilibrium. The estimated frequencies of the
genotypes for the studied genes in the control group did not differ from those earlier reported for Caucasoid women living
in Europe. The T (rs1050450) allele of the GPX1 gene was demonstrated to protect against sporadic breast cancer (OR = 0.74 (95% CI = 0.58−0.94), p = 0.012). Carriers of the genotype combination MnSOD CC + GPX1 CC were found to have a 1.6 times higher risk of sporadic breast cancer compared to the control group (OR = 1.59 (1.05−2.41),
p = 0.0258). The polymorphic loci GSTP1 (rs1695) and MnSOD (rs4880) were not found to be significantly associated with the risk of familial or sporadic breast cancer. 相似文献
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Anber Hassanein Latifa Hamama Karine Loridon Noëlle Dorion 《Plant cell reports》2009,28(10):1521-1530
Direct genetic transformation of mesophyll protoplasts was studied in Pelargonium × hortorum. Calcein and green-fluorescent protein (GFP) gene were used to set up the process. Electroporation (three electric pulses
from a 33-μF capacitor in a 250-V cm−1 electric field) was more efficient than PEG 6000 for membrane permeation, protoplast survival and cell division. Transient
expression of GFP was detected in 33–36% of electroporated protoplasts after 2 days and further in colonies. A protoplast
suspension conductivity of >1,500 μS cm−1 allowed high colony formation and plant regeneration. Stable transformation was obtained using the plasmid FAJ3000 containing
uidA and nptII genes. When selection (50 mg l−1 kanamycin) was achieved 6 weeks after electroporation, regenerated shoots were able to grow and root on 100 mg l−1 kanamycin. The maximum transformation efficiency was 4.5%, based on the number of colonies producing kanamycin-resistant
rooted plants or 0.7% based on the number of cultured protoplasts. Polymerase chain reaction (PCR) analysis on in vitro micropropagated
plants showed that 18 clones out of 20 contained the nptII gene, while the uidA gene was absent. These results were confirmed after PCR analyses of five glasshouse-acclimatized clones. 相似文献
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Zhao-Jun Wei Miao Yu Shun-Ming Tang Yong-Zhu Yi Gui-Yun Hong Shao-Tong Jiang 《Molecular biology reports》2011,38(2):1121-1127
Prothoracicotropic hormone (PTTH) is one of key players in regulation of insect growth, molting, metamorphosis, diapause,
and is expressed specifically in the two pairs of lateral PTTH-producing neurosecretory cells in the brain. Analysis of cis-regulatory elements of the PTTH promoter might elucidate the regulatory mechanism controlling PTTH expression. In this study,
the PTTH gene promoter of Bombyx mori (Bom-PTTH) was cloned and sequenced. The cis-regulatory elements in Bom-PTTH gene promoter were predicted using Matinspector software, including myocyte-specific enhancer
factor 2, pre-B-cell leukemia homeobox 1, TATA box, etc. Transient transfection assays using a series of fragments linked
to the luciferase reporter gene indicated that the fragment spanning −110 to +33 bp of the Bom-PTTH promoter showed high ability
to support reporter gene expression, but the region of +34 to +192 bp and −512 to −111 bp repressed the promoter activity
in the BmN and Bm5 cell lines. Electrophoretic mobility shift assays demonstrated that the nuclear protein could specifically
bind to the region spanning −124 to −6 bp of the Bom-PTTH promoter. Furthermore, we observed that the nuclear protein could
specifically bind to the −59 to −30 bp region of the Bom-PTTH promoter. A classical TATA box, TATATAA, localized at positions
−47 to −41 bp, which is a potential site for interaction with TATA box binding protein (TBP). Mutation of this TATA box resulted
in no distinct binding band. Taken together, TATA box was involved in regulation of PTTH gene expression in B. mori. 相似文献