Meta-analysis and candidate gene mining of low-phosphorus tolerance in maize

Plants with tolerance to low-phosphorus （P） can grow better under Iow-P conditions, and understanding of genetic mechanisms of Iow-P tolerance can not only facilitate identifying relevant genes but also help to develop Iow-P tolerant cultivars. QTL meta-analysis was conducted after a comprehensive review of the reports on O, TL mapping for Iow-P tolerance-related traits in maize. Meta-analysis pro- duced 23 consensus QTL （cQTL）, 17 of which located in similar chromosome regions to those previously reported to influence root traits. Meanwhile, candidate gene mining yielded 215 genes, 22 of which located in the cQTL regions. These 22 genes are homologous to 14 functionally characterized genes that were found to participate in plant Iow-P tolerance, including genes encoding miR399s, Pi transporters and purple acid phosphatases. Four cQTL loci （cQTL2-L cQTLS-3, cQTL6-2, and cQTL10-2） may play important roles for low-P tolerance because each contains more original QTL and has better consistency across previous reports.     

Effects of acetaminophen on hepatic gene expression in mice

Acetaminophen (APAP) is one of the most commonly used drugs for the safe and effective treatment of fever and pain. However, it is a well-established hepatotoxin. The objective of this study was to identify alternation in various genes in liver of mice after administration of low and high doses of APAP. Male C57BL/6J mice received APAP (30 or 300 mg/kg, i.p.). They were sacrificed after 6 hr and 24 hr for assessment of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), total RNA isolation, cDNA microarray analysis and histopathological analysis of liver injury. Low dose of APAP did not cause hepatotoxicity in mice. However, it was toxic at a high dose. Using microarray technology, we selected changed genes more than 1.5 fold. Gene expression changes were recorded even at a low dose treatment with APAP. Six (6) hr after APAP treatment at low dose, 6 genes were up-regulated and 25 genes were down-regulated. However, 24 hr after treatment at low dose 8 genes were up-regulated and 34 genes were down-regulated. 6 hr after of high dose treatment 29 genes were down-regulated and none was up-regulated. A 24 hr treatment with high dose up-regulated 6 genes and down-regulated 18 genes. These expression patterns provide information on high versus low dose mechanisms of APAP toxicity. Gene expression signatures recorded after a nontoxic dose of APAP strongly support the validity of gene expression changes as meaningful markers of hepatotoxicity.     

Yolk polypeptide gene expression in Minute Drosophila females

Minutes have been considered for some time to be mutant at the sites of synthesis of some components of the protein synthetic apparatus. To study the hypothetical relationship between Minutes and suboptimal translation, a group of abundant proteins, the yolk polypeptides, was assayed in outcrossed females bearing M(3)w, M(3)h ^y , or M(1)n mutations. Recently emerged Minute females contained a lower amount of yolk polypeptides, in both ovarian and nonovarian tissues, than their non-Minute sisters. This low level correlated with the lower abundance of cytoplasmic RNA in Minutes compared to control females. By 1 week of age, both M(3)w and their non-Minute sibs contained the same amount of yolk polypeptides and the corresponding mRNA. The double heterozygote, ap ⁴/+;M(3)w/+, did not differ in yolk polypeptide content from control flies. M(3)w females demonstrated reduced fecundity during the period of low yolk polypeptide content but gradually increased egg deposition as yolk polypeptide levels rose. These results suggest that the low protein levels are due to the slower maturation of M(3)w, and not to less efficient translation machinery.This work was supported by the NSERC (Canada) and a Queen's University ARC grant.     

Cellular density regulation of plasminogen gene expression in mouse hepatocytes

The liver produces a variety of proteins including plasminogen. Plasminogen is pro-enzyme that is converted into plasmin by plasminogen activator. Plasmin has a broad substrate spectrum and participates in several biological processes, such as fibrinolysis, tissue remodeling, cell migration, angiogenesis and embryogenesis. In the present study, the regulation of plasminogen expression in mouse hepatocytes was investigated in the primary culture system. Expression level of plasminogen mRNA in the culture at the low cell density condition (0.2 x 10(5) cells / cm(2)) was compared with that at the high cell density condition (1.0 x 10 (5) cells / cm(2)). In the low cell density culture, the expression level of plasminogen mRNA decreased by a time-dependent manner. However, mRNAs for albumin and alpha(2)-antiplasmin were not influenced by the low cell density culture. On the other hand, in the high cell density culture, plasminongen mRNA expressed constantly as well as albumin and alpha(2)-antiplasmin mRNAs. Thus, the decrease in plasminogen mRNA expression could specifically occur when the density of hepatocytes was low. The down-regulation of plasminogen mRNA in the low cell density culture is not observed in the presence of cycloheximide, suggesting that the de novo protein synthesis is required for the regulatory mechanism. These findings indicate that the expression of plasminogen mRNA from hepatocyte is dependent on the cell density and the stimulation by cell-cell contact may be an important factor for the constitutive expression of plasminogen gene in hepatocytes.     

Plant nitrate supply regulates Erwinia amylovora virulence gene expression in Arabidopsis

We showed previously that nitrogen (N) limitation decreases Arabidopsis resistance to Erwinia amylovora (Ea). We show that decreased resistance to bacteria in low N is correlated with lower apoplastic reactive oxygen species (ROS) accumulation and lower jasmonic acid (JA) pathway expression. Consistently, pretreatment with methyl jasmonate (Me-JA) increased the resistance of plants grown under low N. In parallel, we show that in planta titres of a nonvirulent type III secretion system (T3SS)-deficient Ea mutant were lower than those of wildtype Ea in low N, as expected, but surprisingly not in high N. This lack of difference in high N was consistent with the low expression of the T3SS-encoding hrp virulence genes by wildtype Ea in plants grown in high N compared to plants grown in low N. This suggests that expressing its virulence factors in planta could be a major limiting factor for Ea in the nonhost Arabidopsis. To test this hypothesis, we preincubated Ea in an inducing medium that triggers expression of hrp genes in vitro, prior to inoculation. This preincubation strongly enhanced Ea titres in planta, independently of the plant N status, and was correlated to a significant repression of JA-dependent genes. Finally, we identify two clusters of metabolites associated with resistance or with susceptibility to Ea. Altogether, our data showed that high susceptibility of Arabidopsis to Ea, under low N or following preincubation in hrp-inducing medium, is correlated with high expression of the Ea hrp genes in planta and low expression of the JA signalling pathway, and is correlated with the accumulation of specific metabolites.     

c-myc gene expression in human cells is controlled by glucose

The c-myc oncogene is implicated in normal growth and differentiation processes. Human cell lines IM9 and HepG2 stably cultured at "low" glucose concentrations (5.5 mM) show c-myc mRNA levels 3-4 times higher than cells cultured at "high" glucose concentrations (25 nM). D-fructose (a metabolizable exose) substitutes for D-glucose in reducing c-myc expression while 3-ortho-methylglucose (a non metabolizable exose) is uneffective. c-myc expression is up-regulated (by PMA) or down-regulated (by dexamethasone and long-term exposure to FCS) in human cells cultured at "low" glucose but not in cells cultured at "high" glucose. We previously demonstrated that insulin receptor gene expression in human cell lines in enhanced by glucose. Therefore, glucose controls in an opposite way the expression of two genes important in the regulation of eukaryotic cell growth and differentiation.     

Bacterial gene expression at low temperatures

Under suboptimal environmental conditions such as low temperatures, many bacteria have an extended lag phase, altered cell structures, and composition such as a less fluid (more rigid) and leaky cytoplasmic membrane. As a result, cells may die, enter into a starvation mode of metabolism or a physiologically viable but non-culturable (VBNC) state. In the latter state, the amount of gene expression per cell is virtually undetectable. In this article, gene expression under (suboptimal) low temperature conditions in non-psychrophilic environmental bacteria is examined. The pros and cons of some of the molecular methodologies for gene expression analysis are also discussed.     

Identification of RAPD markers linked to a gene governing cadmium uptake in durum wheat.

Temperature sweep gel electrophoresis in combination with random amplified polymorphic DNA analysis was employed to detect two markers for a single gene governing low cadmium uptake in western Canadian durum wheat (Triticum turgidum L. var. durum). Analysis of progeny derived from a cross of the high cadmium accumulating cultivar Kyle by the low cadmium accumulating cultivar Nile resulted in linkage estimates of 4.6 (OPC-20) and 21.2 (UBC-180) cM. The closest marker (OPC-20) was shown to be useful for making low cadmium uptake selections from two other sources of low cadmium, 'Biodur' and 'Hercules'. The marker was further used to determine the genetic basis of resistance in 20 introduced durum wheat lines. Within this diverse range of germplasm the marker was correlated with cadmium contents as expected in all but two cases. Plant breeding selection for low cadmium genotypes is hindered by the high cost of chemical determination of cadmium content. Marker assisted selection for a low cadmium uptake gene offers an effective alternative.     

Nucleofection-based gene targeting in human pre-B cells

Electroporation is a powerful and convenient means for transfection of nonviral vectors into mammalian cells, providing an essential tool for numerous applications including gene targeting via homologous recombination. Recent evidence clearly suggests that high-efficiency gene transfer can be achieved in most cell lines by nucleofection, an electroporation-based transfection method that allows transfected vectors to directly enter the nucleus. In this paper, we analyze the effectiveness of nucleofection for gene targeting using human pre-B cells. For this, we tested 93 different transfection conditions, and found several conditions that gave high (~ 80%) transfection efficiency with low cytotoxicity (~ 70% survival rate). Remarkably, under the optimal nucleofection conditions, the gene-targeting efficiency was ~ 2-5-fold higher than that achieved with conventional electroporation methods. We also found that nucleofection conditions with high transfection efficiency and low cytotoxicity tend to provide high gene-targeting efficiency. Our results provide significant implications for gene targeting, and suggest that nucleofection-based nonviral gene transfer is useful for systematic generation of human gene-knockout cell lines.     

烟草尼古丁去甲基化酶基因及其在育种中的应用

尼古丁通过去甲基化反应生成去甲基尼古丁,后者是潜在的致癌物质亚硝基去甲基尼古丁的合成前体,可能对人体健康产生危害.尼古丁去甲基化酶基因最近被克隆并进化分析,这些基因还作为目标基因应用于低去甲基尼古丁烟草品种选育.该文对近年来该领域研究进展进行综述,并展望该领域未来的研究方向.     

Regulation of alternative oxidase gene expression in soybean

Soybean (Glycine max cv. Stevens) suspension cells were used to investigate the expression of the alternative oxidase (Aox) multigene family. Suspension cells displayed very high rates of cyanide-insensitive respiration, but Aox3 was the only isoform detected in untreated cells. Incubation with antimycin A, citrate, salicylic acid or at low temperature (10 °C) specifically induced the accumulation of the Aox1 isoform. Aox2 was not observed under any conditions in the cells. Increases in Aox1 protein correlated with increases in Aox1 mRNA. Treatment of soybean cotyledons with norflurazon also induced expression of Aox1. Reactive oxygen species (ROS) were detected upon incubation of cells with antimycin, salicylic acid or at low temperature, but not during incubation with citrate. Aox1 induction by citrate, but not by antimycin, was prevented by including the protein kinase inhibitor staurosporine in the medium. The results suggest that multiple pathways exist in soybean to regulate expression of Aox genes and that Aox1 specifically is induced by a variety of stress and metabolic conditions via at least two independent signal transduction pathways.     

Alteration in autophagy gene expression profile correlates with low sperm quality

AimsLow sperm quality, a crucial factor of male infertility, is becoming a public health issue all over the world. In male reproductive system, autophagy plays an important role in maintaining physiological functions. There exist conjectures that disordered autophagy autophagy might be related to low sperm quality. However, there is no evidence can confirm that. This study aims to investigate the association between autophagy-associated genes and low sperm quality.MethodsSperm samples of low sperm quality cases and matched controls were included to select differential expressed genes (DE genes) by autophagy-related functional gene microarray analysis. Then, 104 cases and 250 controls were included to validate the expression of four important autophagy genes (CXCR4, ESR1, PTEN and LC3B). Based on the obtained DE gene, gene Ontology and pathway analyses were conducted.ResultsChip results showed that expression of all 18 DE genes were decreased in the cases compared to the controls (P < 0.05). Expression of ESR1 were verified to be significantly decreased (P < 0.05).ConclusionOur results provided clues with the association among down-regulated expression of autophagy regulating and associated genes and low sperm quality. These findings revealed possible role of impaired autophagy in the mechanism of low sperm quality. Moreover, these may also provide potential targets for the treatment of low sperm quality.     

Growth hormone gene polymorphism associated with selection for milk fat production in lines of cattle

Summary
Fifty-eight calves of both sexes from lines of Red Danish dairy breed selected for high ( n = 36) and low ( n = 22) milk fat production, and 32 heifers from lines of Norwegian Red dairy breed selected for high ( n = 16) and low ( n = 16) milk fat yield were typed for two previously reported restriction fragment length polymorphisms (RFLPs) in the growth hormone gene. The RFLPs are consistent with: (1) an insertion(I)/deletion(D) of approximately 0.9kb in the 3'-region of the growth hormone gene and (2) a polymorphic Msp I(+/−) site in the third intron. A traditional RFLP procedure was used for typing the I/D polymorphism and a polymerase chain reaction (PCR) procedure was developed for typing the Mspl polymorphism. Only the I-MspI(+) and D- Msp I(-) haplotypes were found. In the Red Danish lines the frequency of D- Msp I(-) haplo-type was 0.28 in high line and 0.05 in low line calves, this difference was significant ( P <0.01). The corresponding frequencies in the Norwegian Red lines were 0.09 in the high line and 0.0 in the low line. Attempts to screen for RFLPs in the growth hormone receptor gene and in the insulin-like growth factor-I gene were unsuccessful.     

Polycation liposome-mediated gene transfer in vivo

The polycation liposome (PCL), a recently developed gene transfer system, is simply prepared by a modification of liposomes with cetylated polyethylenimine (PEI), and shows remarkable transgene efficiency with low cytotoxicity. In the present study, we investigated the applicability of PCLs for in vivo gene transfer, since the PCL-mediated transgene efficiency was found to be maintained in the presence of serum. PCLs composed of dioleoylphosphatidylethanolamine (DOPE) with 5 mol% cetyl PEI (PEI average mr. wt. 1800), were superior for transfection to those of dipalmitoylphosphatidylcholine (DPPC) and cholesterol (2:1 as molar ratio) with 5 mol% cetyl PEI in vitro, although the latter PCLs were more efficient for gene transfer in vivo. PCL-DNA complexes were injected into mice via a tail or the portal vein, with the DNA being a plasmid encoding green fluorescent protein (GFP) or luciferase; and the expression was monitored qualitatively or quantitatively, respectively. Tail vein injection resulted in high expression of both GFP and luciferase genes in lung, and portal vein injection resulted in high expression of both genes in the liver. Concerning the gene delivery efficiency, the PCL was found to be superior to PEI or cetyl PEI alone. The optimal conditions for in vivo transfection with PCLs were also examined.     

Polycation liposome-mediated gene transfer in vivo

The polycation liposome (PCL), a recently developed gene transfer system, is simply prepared by a modification of liposomes with cetylated polyethylenimine (PEI), and shows remarkable transgene efficiency with low cytotoxicity. In the present study, we investigated the applicability of PCLs for in vivo gene transfer, since the PCL-mediated transgene efficiency was found to be maintained in the presence of serum. PCLs composed of dioleoylphosphatidylethanolamine (DOPE) with 5 mol% cetyl PEI (PEI average mr. wt. 1800), were superior for transfection to those of dipalmitoylphosphatidylcholine (DPPC) and cholesterol (2:1 as molar ratio) with 5 mol% cetyl PEI in vitro, although the latter PCLs were more efficient for gene transfer in vivo. PCL-DNA complexes were injected into mice via a tail or the portal vein, with the DNA being a plasmid encoding green fluorescent protein (GFP) or luciferase; and the expression was monitored qualitatively or quantitatively, respectively. Tail vein injection resulted in high expression of both GFP and luciferase genes in lung, and portal vein injection resulted in high expression of both genes in the liver. Concerning the gene delivery efficiency, the PCL was found to be superior to PEI or cetyl PEI alone. The optimal conditions for in vivo transfection with PCLs were also examined.