Extensive DNA inversions in the B. fragilis genome control variable gene expression

The obligately anaerobic bacterium Bacteroides fragilis, an opportunistic pathogen and inhabitant of the normal human colonic microbiota, exhibits considerable within-strain phase and antigenic variation of surface components. The complete genome sequence has revealed an unusual breadth (in number and in effect) of DNA inversion events that potentially control expression of many different components, including surface and secreted components, regulatory molecules, and restriction-modification proteins. Invertible promoters of two different types (12 group 1 and 11 group 2) were identified. One group has inversion crossover (fix) sites similar to the hix sites of Salmonella typhimurium. There are also four independent intergenic shufflons that potentially alter the expression and function of varied genes. The composition of the 10 different polysaccharide biosynthesis gene clusters identified (7 with associated invertible promoters) suggests a mechanism of synthesis similar to the O-antigen capsules of Escherichia coli.     

A gene expression map for Caenorhabditis elegans

We have assembled data from Caenorhabditis elegans DNA microarray experiments involving many growth conditions, developmental stages, and varieties of mutants. Co-regulated genes were grouped together and visualized in a three-dimensional expression map that displays correlations of gene expression profiles as distances in two dimensions and gene density in the third dimension. The gene expression map can be used as a gene discovery tool to identify genes that are co-regulated with known sets of genes (such as heat shock, growth control genes, germ line genes, and so forth) or to uncover previously unknown genetic functions (such as genomic instability in males and sperm caused by specific transposons).     

Homeo boxes in the study of development

The body plan of Drosophila is determined to a large extent by homeotic genes, which specify the identity and spatial arrangement of the body segments. Homeotic genes share a characteristic DNA segment, the homeo box, which encodes a defined domain of the homeotic proteins. The homeo domain seems to mediate the binding to specific DNA sequences, whereby the homeotic proteins exert a gene regulatory function. By isolating the normal Antennapedia gene, fusing its protein-coding sequences to an inducible promoter, and reintroducing this fusion gene into the germline of flies, it has been possible to transform head structures into thoracic structures and to alter the body plan in a predicted way. Sequence homologies suggest that similar genetic mechanisms may control development in higher organisms.     

Reversal of oncogenesis by the expression of a major histocompatibility complex class I gene

The classical transplantation antigens (the major histocompatibility complex class I antigens) play a key role in host defense against cells expressing foreign antigens. Several naturally occurring tumors and virally transformed cells show an overall suppression of these surface antigens. Since the class I molecules are required in the presentation of neoantigens on tumor cells to the cytotoxic T lymphocytes, their absence from the cell surface may lead to the escape of these tumors from immunosurveillance. To test this possibility, a functional class I gene was transfected into human adenovirus 12-transformed mouse cells that do not express detectable levels of class I antigens; the transformants were tested for expression of the transfected gene and for changes in oncogenicity. The expression of a single class I gene, introduced by DNA-mediated gene transfer into highly tumorigenic adenovirus 12-transformed cells, was sufficient to abrogate the oncogenicity of these cells. This finding has important implications for the regulation of the malignant phenotype in certain tumors and for the potential modulation of oncogenicity through derepression of the endogenous class I genes.     

Hairpin RNAs and retrotransposon LTRs effect RNAi and chromatin-based gene silencing

The expression of short hairpin RNAs in several organisms silences gene expression by targeted mRNA degradation. This RNA interference (RNAi) pathway can also affect the genome, as DNA methylation arises at loci homologous to the target RNA in plants. We demonstrate in fission yeast that expression of a synthetic hairpin RNA is sufficient to silence the homologous locus in trans and causes the assembly of a patch of silent Swi6 chromatin with cohesin. This requires components of the RNAi machinery and Clr4 histone methyltransferase for small interfering RNA generation. A similar process represses several meiotic genes through nearby retrotransposon long terminal repeats (LTRs). These analyses directly implicate interspersed LTRs in regulating gene expression during cellular differentiation.     

同源重组探针在暂时转化的烟草原生质体内的重组和缺失

通过电激法(electroporation)将同源重组探针pBC 7导入烟草原生质体,再从转化的烟草原生质体中提取DNA,并将其转化到E.coli DH 1细胞.结果在E.codi DH 1细胞中获得了一种新的质粒分子,该质粒含有功能的neo基因,与重组探针pBC 7相比有较大的缺失、倒位,并出现了新的限制酶切部位,重组探针pBC 7的主要特征为含有Tn5的neo基因两个截短的不等部份,只有通过重排才可能产生完整的neo基因,这表明了暂时转化的原生质体在外源DNA整合以前具有这样的基因重排功能。     

DNA methylation and gene function

In most higher organisms, DNA is modified after synthesis by the enzymatic conversion of many cytosine residues to 5-methylcytosine. For several years, control of gene activity by DNA methylation has been recognized as a logically attractive possibility, but experimental support has proved elusive. However, there is now reason to believe, from recent studies, that DNA methylation is a key element in the hierarchy of control mechanisms that govern vertebrate gene function and differentiation.     

人CD46基因启动子的克隆和启动特性研究

为研究人CD46(hCD46)基因启动子的启动特性,研制具有hCD46组织特异性表达特征的转基因小鼠,设计扩增hCD46基因启动子的PCR引物,用HeLa细胞基因组DNA为模板扩增hCD46基因启动子DNA片段,测序结果表明:其序列与GenBank中hCD46完整基因5′端某片段的同源性高达99.9%。用此DNA片段替换表达载体pcD-NA3EGFP中的CMV启动子,且在该片段与EGFP基因之间插入兔β-球蛋白基因的第2内含子RGI。重构的表达载体在2种鼠源细胞CHO和SP2/0中均可启动EGFP表达,表达量与人体内同源组织中hCD46的相似,说明该研究克隆了结构和功能正确的hCD46基因启动子。     

Autogenous regulation of gene expression

A new term, autogenous regulation, is used to describe a phenomenon that is not a new discovery but rather is newly appreciated as a mechanism common to a number of systems in both prokaryotic and eukaryotic organisms. In this mechanism the product of a structural gene regulates expression of the operon in which that structural gene resides. In many (perhaps all) cases, the regulatory gene product has several functions, since it may act not only as a regulatory protein but also as an enzyme, structural protein, or antibody, for example. In a few cases, this protein is the multimeric allosteric enzyme that catalyzes the first step of a metabolic pathway, gearing together the two most important mechanisms for controlling the biosynthesis of metabolites in bacterial cells-feedback inhibition and repression. Autogenous regulation may provide a mechanism for amplification of gene expression (84); for severe and prolonged inactivation of gene expression (85); for buffering the response of structural genes to changes in the environment (45, 52); and for maintaining a constant intracellular concentration of a protein, independent of cell size or growth rate (86). Thus, autogenous regulation provides the cell with means for accomplishing a number of different regulatory tasks, each suited to better satisfying the needs of the organism for its survival.