A hormone response element in the human apolipoprotein CIII (ApoCIII) enhancer is essential for intestinal expression of the ApoA-I and ApoCIII genes and contributes to the hepatic expression of the two linked genes in transgenic mice

We have generated transgenic mice carrying wild-type promoters of the human apolipoprotein A-I (apoA-I)-apoCIII gene cluster or promoters mutated in their hormone response elements. The wild-type cluster directed high levels of apoA-I gene expression in liver and intestine, moderate expression in kidney, and low to minimal expression in other tissues. It also directed high levels of chloramphenicol acetyltransferase (CAT) expression (used as a reporter for the apoCIII gene) in liver, low levels in intestine and kidney, and no expression in other tissues. Mutations in the apoCIII promoter and enhancer abolished the intestinal and renal expression of the apoA-I gene, reduced hepatic apoA-I expression by 80%, and abolished CAT expression in all tissues. A similar pattern of expression was obtained by mutations in the apoCIII enhancer alone. Mutations in the proximal apoA-I promoter reduced by 85% hepatic and intestinal apoA-I expression and did not affect CAT expression. The findings suggest that a hormone response element within the apoCIII enhancer is essential for intestinal and renal expression of apoA-I and apoCIII genes and also enhances hepatic expression. The hormone response elements of the proximal apoA-I promoter or the apoCIII enhancer can promote independently low levels of hepatic and intestinal expression of the apoA-I gene in vivo.     

Different levels of p53 induced either apoptosis or cell cycle arrest in a doxycycline-regulated hepatocellular carcinoma cell line <Emphasis Type="Italic">in vitro</Emphasis>

Induction of p53 gene expression in cancer cells can lead to both cell cycle arrest and apoptosis. To clarify whether the level of p53 expression determines the apoptotic response of hepatocellullar carcinoma (HCC) cells, we assessed the effect of various levels of expression of p53 gene on a p53-deficient HCC cell line, Hep3B, utilizing a doxycycline (Dox)-regulated inducible p53 expression system. Our results showed that apoptosis was induced in HCC cells with high levels of p53 expression. However, lower level of p53 expression induced only cell cycle arrest but not apoptosis. Bax expression was up-regulated following high levels of p53 expression, while bcl-2 expression was not altered by the level of p53 expression. Moreover, p21 expression was observed in both high and low expression of p53. These results suggest the level of p53 expression could determine if the HCC cells would go into cell cycle arrest or apoptosis. Bax may participate, at least in part, in inducing p53-dependent apoptosis and the induction of p21 alone was able to cause cell cycle arrest but not apoptosis.     

用cDNA表达阵列分析遗传性癫痫易感大鼠海马和大脑皮质的基因表达

采用Atlas^TM Rat cDNA Expression Array建立遗传性癫痫易感性P77PMC大鼠和正常对照Wistar大鼠的海马与大脑皮质基因表达谱,用Eagle EyeⅡStill Video System(Stratagene)图象分析仪分析两者基因表达谱差异.结果发现海马和大脑皮质中各有15个差异表达基因.海马组织中,12个基因在P77PMC大鼠中高表达而在正常对照Wistar大鼠中低表达,3个基因在正常对照Wistar大鼠中高表达,而在P77PMC大鼠中低表达;大脑皮质中,13个基因在P77PMC大鼠中高表达,而在正常对照Wistar大鼠中低表达,2个基因在正常对照Wistar大鼠中高表达,而在P77PMC大鼠中低表达. 结果说明,P77PMC大鼠与正常对照Wistar大鼠海马和大脑皮质存在多个差异表达基因,这些差异表达基因可能在癫痫的发生中扮演了重要角色.     

2个表达单元间相互位置和转录方向对表达的影响

目的:研究质粒表达载体中2个表达单元间相互位置和转录方向关系对表达的影响,找出利于2个表达单元表达的优化的相互关系。方法:以全抗体为研究对象,构建了轻重链方向不同的2种相互关系的单质粒表达载体pIRESdhfrA和pIRESdhfrB,转染CHO-dhfr-细胞后,对瞬时表达水平进行了比较。以pIRESdhfrA和pIRESdhfrB为基础,构建了瞬时表达载体pIRESdhfrA-sv40ori和pIRESdhfrB-sv40ori,在COS-7细胞中进行了瞬时表达水平的比较。构建了基于CHO定点细胞系的稳定表达细胞株,对pIRESdhfrA和pIRESdhfrB进行稳定表达水平的比较。结果:在CHO-dhfr-的瞬时表达水平比较中,pIRESdhfrB转染的细胞表达水平是pIRESdhfrA转染细胞的2.18倍。与pIRESdhfrA-sv40ori在COS-7细胞的瞬时表达水平相比,pIRESdhfrB-sv40ori是其表达水平的2.3倍。pIRESdhfrB-FRT构建的CHO定点细胞平均表达水平是pIRESdhfrA-FRT构建的CHO定点细胞的11.6倍。结论:在构建的真核细胞质粒表达载体pIRESdhfr中,2个表达盒的启动子头-头转录方向关系比头-尾方向关系更利于重组蛋白的表达。     

Differential modulation of surfactant protein D under acute and persistent hypoxia in acute lung injury

Hypoxia contributes to the development of fibrosis with epithelial-mesenchymal transition (EMT) via stimulation of hypoxia-inducible factor 1α (HIF-1α) and de novo twist expression. Although hypoxemia is associated with increasing levels of surfactant protein D (SP-D) in acute lung injury (ALI), the longitudinal effects of hypoxia on SP-D expression in lung tissue injury/fibrosis have not been fully evaluated. Here, the involvement of hypoxia and SP-D modulation was evaluated in a model of bleomycin-induced lung injury. We also investigated the molecular mechanisms by which hypoxia might modulate SP-D expression in alveolar cells, by using a doxycycline (Dox)-dependent HIF-1α expression system. Tissue hypoxia and altered SP-D levels were present in bleomycin-induced fibrotic lesions. Acute hypoxia induced SP-D expression, supported by the finding that Dox-induced expression of HIF-1α increased SP-D expression. In contrast, persistent hypoxia repressed SP-D expression coupled with an EMT phenotype and twist expression. Long-term expression of HIF-1α caused SP-D repression with twist expression. Ectopic twist expression repressed SP-D expression. The longitudinal observation of hypoxia and SP-D levels in ALI in vivo was supported by the finding that HIF-1α expression stabilized by acute hypoxia induced increasing SP-D expression in alveolar cells, whereas persistent hypoxia induced de novo twist expression in these cells, causing repression of SP-D and acquisition of an EMT phenotype. Thus this is the first study to demonstrate the molecular mechanisms, in which SP-D expression under acute and persistent hypoxia in acute lung injury might be differentially modulated by stabilized HIF-1α expression and de novo twist expression.     

生产疫苗的植物表达系统

综述了近些年来转基因植物作为疫苗表达系统的新进展,比较了植物表达系统相对其他表达系统的优点,介绍了不同介导方式的植物表达系统,包括农杆菌介导的转化和重组植物病毒表达疫苗,讨论了提高表达效率的载体构建策略,对其应用前景提出了展望。     

Comparative Analysis of Proteome and Transcriptome in Human Hepatocellular Carcinoma using 2D-DIGE and SAGE

Proteome analysis of human hepatocellular carcinoma was conducted using two-dimensional difference gel electrophoresis, and the protein expression profiles were compared to the mRNA expression profiles made from serial analysis of gene expression (SAGE) in identical samples from a single patient. Image-to-image analysis of protein abundances together with protein identification by peptide mass fingerprinting yielded the protein expression profiles. A total of 188 proteins were identified, and the expression profiles of 164 proteins which had the corresponding SAGE data were compared to the mRNA expression profiles. Among them, 40 proteins showed significant differences in the mRNA expression levels between non HCC and HCC. We compared expression changes of proteins with those of mRNAs. We found that the expression tendency of 24 proteins were similar to that of mRNA, whereas 16 proteins showed different or opposite tendency to the mRNA expression.     

Expression of somatostatin receptor mRNAs is regulated in vivo by growth hormone, insulin, and insulin-like growth factor-I in rainbow trout (Oncorhynchus mykiss)

Somatostatins are a diverse family of peptide hormones that regulate various aspects of growth, development, and metabolism through interactions with numerous somatostatin receptor subtypes (SSTRs) on target tissues. In this study, we used rainbow trout to evaluate the effects of growth hormone (GH), insulin (INS), and insulin-like growth factor-I (IGF-I) on the expression of SSTR 1A, 1B and 2 mRNAs. GH regulated the expression of SSTRs in a subtype- and tissue-specific manner. GH reduced SSTR 1A, 1B, and 2 expression in optic tectum, reduced SSTR 1A and 1B expression in pancreas, reduced SSTR 1A expression in liver, and increased hepatic SSTR 1B expression. INS also regulated SSTR expression in a subtype- and tissue-specific manner. INS reduced SSTR 1B expression in optic tectum, increased SSTR 2 expression in pancreas, and increased SSTR 1B and 2 expression in liver. IGF-I generally decreased the expression of all SSTRs. These data indicate that GH, INS, and IGF-I modulate the expression of SSTRs and suggest that independent mechanisms may serve to regulate the various receptor subtypes.     

Effect of mammogenic hormones on the expression of FGF7, FGF10 and their receptor in mouse mammary gland

To investigate the regulation of estrogen, progesterone and prolactin stimulating the development of mammary gland, the Kunming mice were used as experimental animals in this study. Through the experiment in vitro, the effect of mammogenic hormones were systematically investigated on expression of FGF7 and FGF10 and their receptor in different periods. The results are as follows: in mammary glands of mice, 17 beta-estradiol increased the expression of FGF7; progesterone did not affect the expression of FGF7; prolactin up-regulated the expression of FGF7 significantly in pregnancy and lactation. 17 beta-estradiol increased the expression of FGF10; progesterone and prolactin reduced the expression of FGF10 significantly in virgin; prolactin significantly increased the expression of FGF10 in pregnancy. When 17 beta-estradiol in the body was in relatively high proportion, it would lower the expression of KGFR; while 17 beta-estradiol in the body was in relatively low proportion, it would increase the expression of KGFR. Low concentration of progesterone increased the expression of KGFR and high progesterone did not affect the expression of KGFR. Prolactin increased the expression of KGFR significantly in pregnancy and lactation.     

抑制Dicer基因对shRNA功能发挥的影响

本文将Dicer基因的RNA酶III结构域作为靶区,设计并构建了两个抗Dicer基因的小发夹样RNA(shRNA)表达载体,将其转染2215、结肠癌TC细胞和基因组中整合有绿色荧光蛋白基因(GFP)的HepG2A9细胞,通过RT-PCR评价RNA干扰抑制Dicer基因表达的效率;当HepG2A9细胞Dicer基因表达被上述RNA干扰抑制时,再转染抗GFP的shRNA表达载体,通过RT-PCR和荧光显微镜观察GFP表达水平。结果显示,在不同细胞系中,这两个抗Dicer基因shRNA表达载体,均能明显抑制Dicer基因的表达;当Dicer基因受抑时,后续转染抗GFP的shRNA表达载体不能有效抑制GFP的表达。结果表明,抗Dicer基因shRNA表达载体,能够明显抑制Dicer基因的表达;shRNA表达载体的功能发挥需要Dicer酶的直接参与。     

含p53保守结合位点的microRNA表达载体的构建及应用

目的:构建含p53保守结合位点的microRNA(miRNA)表达载体,促进相关miRNA在具有野生型p53蛋白细胞中的高效表达。方法:改构miRNA表达载体pCMV-miR,在其多克隆位点前插入p53保守结合位点,分别将miR-138、miR-34a和miR-21前体序列pre-miR-138、pre-miR-34a和pre-miR-21插入上述改构的载体pCMV/p53-miR,将构建的pCMV/p53-miR-138、pCMV/p53-miR-34a和pCMV/p53-miR-21表达载体转染具有野生型p53的HeLa细胞和不表达p53的H1299细胞,分析p53对上述miRNA表达调控的影响。结果:转染改构的miRNA表达载体后,HeLa细胞中miR-138、miR-34a和miR-21的表达水平明显提高,它们对应的已知靶基因Cyclin D3、CDK2和PTEN的表达同时被显著下调。结论:在p53转录调控作用下,具有p53保守结合位点的miRNA表达载体能够更加有效地提高miRNA的表达水平;构建的载体不但可用于促进相关miRNA的表达,也能用于miRNA是否受p53调控的检测。     

Positive selection for elevated gene expression noise in yeast

It is well known that the expression noise is lessened by natural selection for genes that are important for cell growth or are sensitive to dosage. In theory, expression noise can also be elevated by natural selection when noisy gene expression is advantageous. Here we analyze yeast genome‐wide gene expression noise data and show that plasma‐membrane transporters show significantly elevated expression noise after controlling all confounding factors. We propose a model that explains why and under what conditions elevated expression noise may be beneficial and subject to positive selection. Our model predicts and the simulation confirms that, under certain conditions, expression noise also increases the evolvability of gene expression by promoting the fixation of favorable expression level‐altering mutations. Indeed, yeast genes with higher noise show greater between‐strain and between‐species divergences in expression, even when all confounding factors are excluded. Together, our theoretical model and empirical results suggest that, for yeast genes such as plasma‐membrane transporters, elevated expression noise is advantageous, is subject to positive selection, and is a facilitator of adaptive gene expression evolution.     

Evolution of duplicate gene expression in polyploid and hybrid plants

Allopolyploidy is a prominent mode of speciation in flowering plants. On allopolyploidy, genomic changes can take place, including chromosomal rearrangement and changes in gene expression; these processes continue over evolutionary time. Recent studies of gene expression in polyploid and hybrid plants, reviewed here, have examined expression in natural polyploids and synthetic neopolyploids as well as in diploid and F(1) hybrids. Considerable changes in gene expression have been observed in allopolyploids, including up- or downregulation of expression in the polyploids compared with their parents, unequal expression of duplicated genes, and silencing of one copy. Genes in a variety of functional categories show altered expression, and the patterns vary considerably by gene. Some changes seem to be stochastic, whereas others are repeatable. Gene expression changes can be organ specific. Reciprocal silencing of duplicates in different organs has been observed, suggesting subfunctionalization and long-term retention of duplicates. It has become clear that hybridization has a much greater effect than chromosome doubling on gene expression in allopolyploids. Diploid and triploid F(1) hybrids can show alterations of expression levels compared with their parents. Parent-of-origin effects on gene expression have been examined, and loss of gene imprinting has been shown. Some gene expression changes in polyploids and hybrids can be correlated with phenotypic effects. Demonstrated mechanisms of gene expression changes include DNA methylation, histone modifications, and antisense RNA. Several hypotheses have been proposed for why gene expression is altered in allopolyploids and hybrids.     

5-Hydroxytryptamine 2A receptor signaling cascade modulates adiponectin and plasminogen activator inhibitor 1 expression in adipose tissue

Knowledge of the regulatory factors associated with down-regulation of adiponectin gene expression and up-regulation of PAI-1 gene expression is crucial to understand the pathophysiological basis of obesity and metabolic diseases, and could establish new treatment strategies for these conditions. We showed that expression of 5-HT(2A) receptors was up-regulated in hypertrophic 3T3-L1 adipocytes, which exhibited decreased expression of adiponectin and increased expression of PAI-1. 5-HT(2A) receptor antagonists and suppression of 5-HT(2A) receptor gene expression enhanced adiponectin expression. Activation of Gq negatively regulated adiponectin expression, and inhibition of mitogen-activated protein kinase reversed the Gq-induced effect. Moreover, the 5-HT(2A) receptor blockade reduced PAI-1 expression. These findings indicate that antagonism of 5-HT(2A) receptors in adipocytes could improve the obesity-linked decreases in adiponectin expression and increases in PAI-1 expression.     

Thymidine phoshorylase expression in breast cancer: the prognostic significance and its association with other angiogenesis related proteins and extracellular matrix components

Thymidine phosphorylase (TP)/platelet-derived endothelial cell growth factor, stimulates chemataxis of endothelial cells and is involved in the angiogenesis of human solid tumours. In this study we investigated tissue sections from 93 breast carcinomas for the immunohistochemical expression of thymidine phosphorylase protein and in relationship to several clinicopathological parameters. The possible relationship to tumour neovascularization, VEGF expression, extracellular matrix components (tenascin, fibronectin, collagen type IV and laminin) and cathepsin D was also estimated. Nuclear and/or cytoplasmic TP expression was observed in tumour cells. Immunoreactivity was also often present in the stroma, endothelium and tumour-associated macrophages. High cytoplasmic TP expression, was observed in 35.5%, moderate in 30.1%, mild in 18.3%, while 16.1% of the cases were negative for TP expression. Moderate and high nuclear TP expression was observed in 30.1% of the tumours, low in 43%, while 26.9% did not show nuclear TP expression. High tumour stroma TP expression was expressed in 23.7% of the cases, moderate in 21.5%, mild in 45.2%, while 9.7% did not show stromal TP expression. TP expression did not correlate with the conventional clinicopathological features as well as with the microvessel density and the VEGF expression. Patients with high levels of tumour cell TP expression were significantly associated with a favorable outcome in univariate method of analysis. A positive correlation of TP expression with Cathepsin D expression was noticed. In addition, tumour cell TP expression was correlated with the extracellular matrix component tenascin, while stromal cell TP expression was correlated with the growth fraction of the tumour. Our data suggests that TP expression does not seem to affect directly the neovasculatur of breast carcinoma, although it seems to be implicated in the remodeling of breast cancer tissue, through the interaction with other extracellular matrix components or proteolytic enzymes. In addition, tumour cell TP expression could be considered as a prognostic indicator of breast cancer patients.     

Regulation of type II collagen expression by cyclin-dependent kinase 6, cyclin D1, and p21 in articular chondrocytes

This study examined whether cell cycle regulatory proteins, such as cyclin-dependent kinases (CDKs), cyclins, and CDK inhibitors, regulate type II collagen expression and mediate interlukin-1 (IL-1beta)-induced suppression of type II collagen expression in articular chondrocytes. IL-1beta inhibited type II collagen expression, but activated CDK6. Ectopic expression of CDK2 did not alter type II collagen expression. However, overexpression of CDK6 inhibited type II collagen expression, whereas inhibition of CDK6 activity blocked IL-1beta-induced suppression of type II collagen expression. IL-1beta upregulated the expression of cyclin D1, which is known to activate CDK6. In turn, overexpression of cyclin D1 suppressed type II collagen expression. In contrast to cyclin D1, IL-1beta triggered down-regulation of the CDK inhibitor, p21. Overexpression of p21 blocked IL-1beta- or CDK6-induced suppression of type II collagen expression. Our results collectively indicate that CDK6/cyclin D1/p21 complex regulates type II collagen expression in articular chondrocytes.     

Databases of free expression

The rapid development of microarray technologies has led to a similar progression in gene expression analysis methods, gene expression applications, and gene expression databases. Public gene expression databases enable any researcher to examine expression of their favorite genes across a wide variety of samples, download sample data for development of new analysis methods, or answer broad questions about gene expression regulation, among other applications. A wide variety of public gene expression databases exist, and they vary in their content, analysis capabilities, and ease of use. This review highlights the current features and describes examples of two broad categories of mammalian microarray databases: tissue gene expression databases and data warehouses.