Intron length variation of the Adh gene in Brachyscome (Asteraceae)

Eighteen polymerase chain reaction (PCR) products of the partial sequence of the Adh (alcohol dehydrogenase) gene from 10 Brachyscome species were sequenced and compared. These products contained the 5 three fourths of exon 4 and whole sequences of intron 3. They varied extensively in length due to the differences in length of intron 3. A total of 10 long insertions were flanked by direct repeats of 5 to 12 bp sequences, indicating inserted elements. These inserted elements were classified into the following five categories based on nucleotide sequence characteristics and length; (1) a region homologous to that of 5S RNA genes (5S DNA), (2) A-rich structure at the 3 end-like short interspersed elements (SINEs) in animals, (3) a sequence of 280 bp with no characteristic features, (4) a sequence of 125 bp with no characteristic features, (5) termini of 11 bp inverted repeats flanked by 5 bp sequence of direct repeats characteristics of a transposon.     

Evidence for a host role in regulating the activity of transposable elements in Drosophila melanogaster: the case of the persistent instability of Bari 1 elements in Charolles stock

In most reports in which the activity of numerous Drosophila transposon families has been studied, only a subset of the families tested appears mobile. A comparison of these data shows that there are no transposons inherently more unstable than others and suggests that host factors regulate the activity of transposable elements. Consistent with this conclusion are the properties of Bari 1 elements, which are the only ones of the 14 families tested to be unstable in Charolles stock. Instability is persistent over 53 generations and appears to affect recurrent insertion sites. This revised version was published online in August 2006 with corrections to the Cover Date.     

Transposable Elements Cross Kingdom Boundaries and Contribute to Inflammation and Ageing

The de-repression of transposable elements (TEs) in mammalian genomes is thought to contribute to genome instability, inflammation, and ageing, yet is viewed as a cell-autonomous event. In contrast to mammalian cells, prokaryotes constantly exchange genetic material through TEs, crossing both cell and species barriers, contributing to rapid microbial evolution and diversity in complex communities such as the mammalian gut. Here, it is proposed that TEs released from prokaryotes in the microbiome or from pathogenic infections regularly cross the kingdom barrier to the somatic cells of their eukaryotic hosts. It is proposed this horizontal transfer of TEs from microbe to host is a stochastic, ongoing catalyst of genome destabilization, resulting in structural and epigenetic variations, and activation of well-evolved host defense mechanisms contributing to inflammation, senescence, and biological ageing. It is proposed that innate immunity pathways defend against the horizontal acquisition of microbial TEs, and that activation of this pathway during horizontal transposon transfer promotes chronic inflammation during ageing. Finally, it is suggested that horizontal acquisition of prokaryotic TEs into mammalian genomes has been masked and subsequently under-reported due to flaws in current sequencing pipelines, and new strategies to uncover these events are proposed.     

利用转座子构建靶向侵袭性抗肿瘤工程菌

转座子是一类在基因组上可以自由跳跃的移动序列,同时也是对微生物进行基因修饰和插入突变的有效工具,但尚未见有利用转座子导入革兰氏阴性菌E.coli Nissle1917菌株的报道.本研究通过构建p R6K转座载体,对肠道益生菌E.coli Nissle1917菌株进行了转座插入诱变,将假结核耶尔森菌的侵袭素基因inv和单核细胞增多性李斯特菌的溶血素基因hly随机整合至E.coli Nissle1917菌株的染色体上,从而使非致病性大肠杆菌E.coli Nissle1917获得侵袭哺乳动物细胞的能力.通过细胞体外侵袭实验发现,本研究所构建的工程菌对B16,HCT-116等肿瘤细胞有较好的侵袭活性,同时与抗肿瘤蛋白Azurin一起作用B16细胞,抗肿瘤效果显著增强,为进一步运用以大肠杆菌E.coli Nissle1917作为DNA疫苗或者基因治疗的载体开辟了新的技术途径.     

Recent evolution of the human pathogen Cryptococcus neoformans by intervarietal transfer of a 14-gene fragment

The availability of the whole-genome sequence from the 2 known varieties of the human pathogenic fungus Cryptococcus neoformans provides an opportunity to study the relative contribution of divergence and introgression during the process of speciation in a genetically tractable organism. At the genomic level, these varieties are nearly completely syntenic, share approximately 85-90% nucleotide identity, and are believed to have diverged approximately 18 MYA. Via a comparative genomic approach, we identified a 14-gene region (approximately 40 kb) that is nearly identical between the 2 varieties that resulted from a nonreciprocal transfer event from var. grubii to var. neoformans approximately 2 MYA. The majority of clinical and environmental var. neoformans strains from around the world contain this sequence obtained from var. grubii. This introgression event likely occurred via an incomplete intervarietal sexual cycle, creating a hybrid intermediate where mobile elements common to both lineages mediated the exchange. The subsequent duplication in laboratory strains of a fragment of this same genomic region supports evolutionary theories that instabilities in subtelomeric regions promote adaptive evolution through gene amplification and subsequent adaptation. Along with a more ancient predicted transfer event in C. neoformans and a recently reported example from Saccharomyces cerevisiae, these data indicate that DNA exchange between closely related sympatric varieties or species may be a recurrent theme in the evolution of fungal species. It further suggests that although evolutionary divergence is the primary force driving speciation, rare introgression events also play a potentially important role.     

Are we Genomic Mosaics? Variations of the Genome of Somatic Cells can Contribute to Diversify our Phenotypes

Theoretical and experimental evidences support the hypothesis that the genomes and the epigenomes may be different in the somatic cells of complex organisms. In the genome, the differences range from single base substitutions to chromosome number; in the epigenome, they entail multiple postsynthetic modifications of the chromatin. Somatic genome variations (SGV) may accumulate during development in response both to genetic programs, which may differ from tissue to tissue, and to environmental stimuli, which are often undetected and generally irreproducible. SGV may jeopardize physiological cellular functions, but also create novel coding and regulatory sequences, to be exposed to intraorganismal Darwinian selection. Genomes acknowledged as comparatively poor in genes, such as humans', could thus increase their pristine informational endowment. A better understanding of SGV will contribute to basic issues such as the "nature vs nurture" dualism and the inheritance of acquired characters. On the applied side, they may explain the low yield of cloning via somatic cell nuclear transfer, provide clues to some of the problems associated with transdifferentiation, and interfere with individual DNA analysis. SGV may be unique in the different cells types and in the different developmental stages, and thus explain the several hundred gaps persisting in the human genomes "completed" so far. They may compound the variations associated to our epigenomes and make of each of us an "(epi)genomic" mosaic. An ensuing paradigm is the possibility that a single genome (the ephemeral one assembled at fertilization) has the capacity to generate several different brains in response to different environments.     

Mycobacteria: Bugs and bugbears (Two steps forward and one step back)

The use of molecular techniques to study the mycobacteria has advanced greatly since the first genomic libraries of Mycobacterium tuberculosis and M. leprae were constructed in 1985. However, there are still pitfalls for the unwary. Most of the problems associated with the use of molecular techniques to study mycobacteria can be related to one of the following problems: slow growth rate causing problems with contamination; the formation of macroscopic clumps when grown in culture; resistance to standard chemical lysis procedures; the requirement for containment facilities for pathogenic species; the lack of suitable genetic vectors; and the problems of spontaneous antibiotic resistance. Despite these problems, considerable progress has been made and standard techniques have been developed for the preparation of protein, nucleic acids (DNA and RNA) and cell wall components, chemical and transposon mutagenesis and gene replacement methods, the use of reporter genes and expression vectors, and improved detection and drug sensitivity testing.     

SIMILAR UNSTABLE MUTATIONS IN THREE SPECIES OF GRACILARIA (RHODOPHYTA)1

Unstable mutants with similar variegated pigmentation were genetically characterized in the red algae. Gracilaria tikvahiae (McLachlan), G. foliifera (Forsk.) Børg. and. G. sjoestedtii (Kylin). All three mutants were green plants with flecks of red tissue where cells had reverted to wild type. The mutant green phenotypes were all recessive, and their genetic behavior in crosses indicated that each was the result of a single, unstable, nuclear gene. Wild-type revertant tissue was stable one it arose. Revertant plants obtained from spores and revertant fronds taken from variegated plants could not be distinguished from the normal wild type, either phenotypically or genetically. Reversion to wild type occurred during all phases of the life cycle. In crosses between the mutants and wild type, most of the F₁ tetrasporophytes were heterozygous wild-type plants, an observation consistent with the recessive nature of the mutations; however, a low frequency of homozygous unstable-green F₁ tetrasporophytes was also obttained from these crosses. The molecular basis of neither the mutant instability, i.e. the reversion to wild type, nor of the process producing the unstable green F₁ tetrasporophytes can yet be deduced, but the phenotype of the plants and genetic results suggest the involvement of transposable genetic elements.     

DNA binding domains in Tn3 transposase

Summary Various segments of Tn3 transposase were fused individually to -galactosidase, and the resulting fusion proteins were examined for their DNA binding ability by a nitrocellulose filter binding assay. Analyses of a series of the fusion proteins revealed that the N-terminal segment of the transposase (amino acid positions 1–242; the transposase gene encodes 1004 residues in all) had specific DNA binding ability for the 38 bp terminal inverted repeat (IR) sequence, and the central segment (amino acid positions 243–632) had non-specific DNA binding ability. Further analyses of each of the two regions revealed that the N-terminal segment could be divided into at least two subsegments (amino acid positions 1–86 and 87–242), neither of which had specific DNA binding ability, but which both possessed nonspecific DNA binding ability. The central segment included two subsegments (amino acid positions 243–289 and 439–505) with non-specific DNA binding ability. These results and other observations suggest that Tn3 transposase has several domains including those responsible for non-specific DNA binding, and a combination of two or more domains gives rise to specific DNA binding activity. The C-terminal segment of the transposase (amino acid positions 633-1004), which is very well conserved among transposases encoded by Tn3 family transposons, had no DNA binding ability. This segment may represent the main part of the catalytic domain responsible for the initiation step of transposition.