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PubMed2006

Positive selection of yeast nonhomologous end-joining genes and a retrotransposon conflict hypothesis.

Transposable elements have clearly played a major role in shaping both the size and organization of eukaryotic genomes. However, the evolution of essential genes in core biological processes may also have been shaped by coevolution with these elements. This would be predicted to occur in instances where host proteins are either hijacked for use by mobile elements or recruited to defend against them. To detect such cases, we have used the Saccharomyces cerevisiae-Saccharomyces paradoxus sibling species pair to identify genes that have evolved under positive selection. We identify 72 such genes, which participate in a variety of biological processes but are enriched for genes involved in meiosis and DNA repair by nonhomologous end-joining (NHEJ). We confirm the signature of positive selection acting on NHEJ genes using orthologous sequences from all seven Saccharomyces sensu stricto species. Previous studies have found altered rates of Ty retrotransposition when these NHEJ genes are disrupted. We propose that the evolution of these repair proteins is likely to have been shaped by their interactions with Ty elements. Antagonistic pleiotropy, where critical genes like those involved in DNA repair are also subject to selective pressures imposed by mobile elements, could favor alleles that might be otherwise deleterious for their normal roles related to genome stability.

Base Sequence↗
PubMed2001

Integration of matrix-assisted laser desorption ionization-time of flight mass spectrometry and molecular cloning for the identification and functional characterization of mobile ortho-halobenzoate oxygenase genes in Pseudomonas aeruginosa strain JB2.

Protein mass spectrometry and molecular cloning techniques were used to identify and characterize mobile o-halobenzoate oxygenase genes in Pseudomonas aeruginosa strain JB2 and Pseudomonas huttiensis strain D1. Proteins induced in strains JB2 and D1 by growth on 2-chlorobenzoate (2-CBa) were extracted from sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels and analyzed by matrix-assisted laser desorption ionization-time of flight mass spectrometry. Two bands gave significant matches to OhbB and OhbA, which have been reported to be the alpha and beta subunits, respectively, of an ortho-1,2-halobenzoate dioxygenase of P. aeruginosa strain 142 (T. V. Tsoi, E. G. Plotnikova, J. R. Cole, W. F. Guerin, M. Bagdasarian, and J. M. Tiedje, Appl. Environ. Microbiol. 65:2151-2162, 1999). PCR and Southern hybridization experiments confirmed that ohbAB were present in strain JB2 and were transferred from strain JB2 to strain D1. While the sequences of ohbA from strains JB2, D1, and 142 were identical, the sequences of ohbB from strains JB2 and D1 were identical to each other but differed slightly from that of strain 142. PCR analyses and Southern hybridization analyses indicated that ohbAB were conserved in strains JB2 and D1 and in strain 142 but that the regions adjoining these genes were divergent. Expression of ohbAB in Escherichia coli resulted in conversion of o-chlorobenzoates to the corresponding (chloro)catechols with the following apparent affinity: 2-CBa approximately 2,5-dichlorobenzoate > 2,3,5-trichlorobenzoate > 2,4-dichlorobenzoate. The activity of OhbAB(JB2) appeared to differ from that reported for OhbAB(142) primarily in that a chlorine in the para position posed a greater impediment to catalysis with the former. Hybridization analysis of spontaneous 2-CBa(-) mutants of strains JB2 and D1 verified that ohbAB were lost along with the genes, suggesting that all of the genes may be contained in the same mobile element. Strains JB2 and 142 originated from California and Russia, respectively. Thus, ohbAB and/or the mobile element on which they are carried may have a global distribution.

Blotting, Southern↗
PubMed1992

Structural organization and diversification of Y-linked sequences comprising Su(Ste) genes in Drosophila melanogaster.

Expression of the X-linked repeated Stellate (Ste) genes, which code for a protein with 38% similarity to the beta-subunit of casein kinase II, is suppressed by the Su(Ste) locus on the Y chromosome. The structure and evolution of the Y-linked repeats in the region of the Su(Ste) locus were studied. The 2800 bp repeats consist of three main elements: the region of homology to the Ste genes, an adjacent AT-rich, Y-specific segment, and mobile element 1360 inserted in the Ste sequence. Amplification of repeats was followed by point mutations, deletions, and insertions of mobile elements. DNA sequencing shows that these repeats may be considered as Ste pseudogenes or as damaged variants of a putative gene(s) encoding a protein quite different from the Ste protein as a result of an alternative splicing pattern. A comparison of 5 variants of the Y-Su(Ste) repeats shows a number of recombination events between amplified and diverged sequences that could be due to either multiple unequal mitotic sister-chromatid exchanges or to gene conversion. It is a first demonstration on a molecular level of these processes occurring in heterochromatic non-rDNA tandemly organized sequences in an eukaryotic genome.

Animals↗
PubMed2006

Gamma radiation increases endonuclease-dependent L1 retrotransposition in a cultured cell assay.

Long Interspersed Elements (LINE-1s, L1s) are the most active mobile elements in the human genome and account for a significant fraction of its mass. The propagation of L1 in the human genome requires disruption and repair of DNA at the site of integration. As Barbara McClintock first hypothesized, genotoxic stress may contribute to the mobilization of transposable elements, and conversely, element mobility may contribute to genotoxic stress. We tested the ability of genotoxic agents to increase L1 retrotransposition in a cultured cell assay. We observed that cells exposed to gamma radiation exhibited increased levels of L1 retrotransposition. The L1 retrotransposition frequency was proportional to the number of phosphorylated H2AX foci, an indicator of genotoxic stress. To explore the role of the L1 endonuclease in this context, endonuclease-deficient tagged L1 constructs were produced and tested for their activity in irradiated cells. The activity of the endonuclease-deficient L1 was very low in irradiated cells, suggesting that most L1 insertions in irradiated cells still use the L1 endonuclease. Consistent with this interpretation, DNA sequences that flank L1 insertions in irradiated cells harbored target site duplications. These results suggest that increased L1 retrotransposition in irradiated cells is endonuclease dependent. The mobilization of L1 in irradiated cells potentially contributes to genomic instability and could be a driving force for secondary mutations in patients undergoing radiation therapy.

Aminoglycosides↗
PubMed2006

Positive selection on transposase genes of insertion sequences in the Crocosphaera watsonii genome.

Insertion sequences (ISs) are mobile elements that are commonly found in bacterial genomes. Here, the structural and functional diversity of these mobile elements in the genome of the cyanobacterium Crocosphaera watsonii WH8501 is analyzed. The number, distribution, and diversity of nucleotide and amino acid stretches with similarity to the transposase gene of this IS family suggested that this genome harbors many functional as well as truncated IS fragments. The selection pressure acting on full-length transposase open reading frames of these ISs suggested (i) the occurrence of positive selection and (ii) the presence of one or more positively selected codons. These results were obtained using three data sets of transposase genes from the same IS family that were collected based on the level of amino acid similarity, the presence of an inverted repeat, and the number of sequences in the data sets. Neither recombination nor ribosomal frameshifting, which may interfere with the selection analyses, appeared to be important forces in the transposase gene family. Some positively selected codons were located in a conserved domain, suggesting that these residues are functionally important. The finding that this type of selection acts on IS-carried genes is intriguing, because although ISs have been associated with the adaptation of the bacterial host to new environments, this has typically been attributed to transposition or transformation, thus involving different genomic locations. Intragenic adaptation of IS-carried genes identified here may constitute a novel mechanism associated with bacterial diversification and adaptation.

Adaptation, Biological↗
PubMed1998

Mobilization of heavy metals and arsenic in polluted wetland soils and its dependence on dissolved organic matter.

The wetland soils of the Mulde river in the industrial district of Bitterfeld-Wolfen (Germany) are highly contaminated with heavy metals and arsenic. We studied the mobility of accumulated heavy metals and arsenic and the influence of dissolved organic matter (DOM) on element mobility. Undisturbed soil cores were taken from five different sites to represent a wide range of heavy-metal contamination, soil properties and dissolved organic carbon (DOC) concentrations. The acid-soluble concentrations (mostly equal to the total content) were up to 1100 mg kg-1 for Zn, 800 mg kg-1 for Cr, 364 mg kg-1 for Cu, 265 mg kg-1 for As and 37 mg kg-1 for Hg, depending on the sampling site. Percolation experiments using small lysimeters with undisturbed topsoil cores illustrated a considerable mobilization of Zn, Cd, Cu, Cr and Hg, depending on soil properties. Up to 80 micrograms l-1 Cd, 8 mg 1-1 Zn, 130 micrograms l-1 Cr, 160 micrograms l-1 Cu and 7 micrograms l-1 Hg were detected in the soil percolates. Arsenic mobilization was low. The concentration of Cr, Hg, Cu and As in the soil percolates was positively correlated with DOM. Besides the element content (mobile or acid-soluble), soil pH and soil characteristics describing the soil potential for heavy-metal adsorption (clay, oxides, cation exchange capacity), the DOC concentration in the soil solution should be known to access the potential mobilization of Hg, Cr, Cu and As. In contrast, Cd and Zn mobilization depends on soil pH and mobile element content, but not on DOM. Additional studies on two soil profiles (down to 1.5 m) confirmed the translocation of heavy metals from the highly contaminated topsoil into deeper soil horizons and into the groundwater and the influence of DOM as revealed with the percolation experiment. Our results also showed that DOM is of minor importance on the mobilization of heavy metals in soils with a low soil pH (< 4.5).

Adsorption↗
PubMed2006

Identification and characterization of novel polymorphic LINE-1 insertions through comparison of two human genome sequence assemblies.

Mobile elements represent a relatively new class of markers for the study of human evolution. Long interspersed elements (LINEs) belong to a group of retrotransposons comprising approximately 21% of the human genome. Young LINE-1 (L1) elements that have integrated recently into the human genome can be polymorphic for insertion presence/absence in different human populations at particular chromosomal locations. To identify putative novel L1 insertion polymorphisms, we computationally compared two draft assemblies of the whole human genome (Public and Celera Human Genome assemblies). We identified a total of 148 potential polymorphic L1 insertion loci, among which 73 were candidates for novel polymorphic loci. Based on additional analyses we selected 34 loci for further experimental studies. PCR-based assays and DNA sequence analysis were performed for these 34 loci in 80 unrelated individuals from four diverse human populations: African-American, Asian, Caucasian, and South American. All but two of the selected loci were confirmed as polymorphic in our human population panel. Approximately 47% of the analyzed loci integrated into other repetitive elements, most commonly older L1s. One of the insertions was accompanied by a BC200 sequence. Collectively, these mobile elements represent a valuable source of genomic polymorphism for the study of human population genetics. Our results also suggest that the exhaustive identification of L1 insertion polymorphisms is far from complete, and new whole genome sequences are valuable sources for finding novel retrotransposon insertion polymorphisms.

Alleles↗
PubMed2004

IHF is the limiting host factor in transposition of Pseudomonas putida transposon Tn4652 in stationary phase.

Transpositional activity of mobile elements is not constant. Conditional regulation of host factors involved in transposition may severely change the activity of mobile elements. We have demonstrated previously that transposition of Tn4652 in Pseudomonas putida is a stationary phase-specific event, which requires functional sigma S (Ilves et al., 2001, J Bacteriol 183: 5445-5448). We hypothesized that integration host factor (IHF), the concentration of which is increased in starving P. putida, might contribute to the transposition of Tn4652 as well. Here, we demonstrate that transposition of Tn4652 in stationary phase P. putida is essentially limited by the amount of IHF. No transposition of Tn4652 occurs in a P. putida ihfA-defective strain. Moreover, overexpression of IHF results in significant enhancement of transposition compared with the wild-type strain. This indicates that the amount of IHF is a bottleneck in Tn4652 transposition. Gel mobility shift and DNase I footprinting studies revealed that IHF is necessary for the binding of transposase to both transposon ends. In vitro, transposase can bind to inverted repeats of transposon only after the binding of IHF. The results obtained in this study indicate that, besides sigma S, IHF is another host factor that is implicated in the elevation of transposition in stationary phase.

DNA Footprinting↗
PubMed2002

A new, expressed multigene family containing a hot spot for insertion of retroelements is associated with polymorphic subtelomeric regions of Trypanosoma brucei.

We describe a novel gene family that forms clusters in subtelomeric regions of Trypanosoma brucei chromosomes and partially accounts for the observed clustering of retrotransposons. The ingi and ribosomal inserted mobile element (RIME) non-LTR retrotransposons share 250 bp at both extremities and are the most abundant putatively mobile elements, with about 500 copies per haploid genome. From cDNA clones and subsequently in the T. brucei genomic DNA databases, we identified 52 homologous gene and pseudogene sequences, 16 of which contain a RIME and/or ingi retrotransposon inserted at exactly the same relative position. Here these genes are called the RHS family, for retrotransposon hot spot. Comparison of the protein sequences encoded by RHS genes (21 copies) and pseudogenes (24 copies) revealed a conserved central region containing an ATP/GTP-binding motif and the RIME/ingi insertion site. The RHS proteins share between 13 and 96% identity, and six subfamilies, RHS1 to RHS6, can be defined on the basis of their divergent C-terminal domains. Immunofluorescence and Western blot analyses using RHS subfamily-specific immune sera show that RHS proteins are constitutively expressed and occur mainly in the nucleus. Analysis of Genome Survey Sequence databases indicated that the Trypanosoma brucei diploid genome contains about 280 RHS (pseudo)genes. Among the 52 identified RHS (pseudo)genes, 48 copies are in three RHS clusters located in subtelomeric regions of chromosomes Ia and II and adjacent to the active bloodstream form expression site in T. brucei strain TREU927/4 GUTat10.1. RHS genes comprise the remaining sequence of the size-polymorphic "repetitive region" described for T. brucei chromosome I, and a homologous gene family is present in the Trypanosoma cruzi genome.

Amino Acid Sequence↗
PubMed1999

Analysis of two cosmid clones from chromosome 4 of Drosophila melanogaster reveals two new genes amid an unusual arrangement of repeated sequences.

Chromosome 4 from Drosophila melanogaster has several unusual features that distinguish it from the other chromosomes. These include a diffuse appearance in salivary gland polytene chromosomes, an absence of recombination, and the variegated expression of P-element transgenes. As part of a larger project to understand these properties, we are assembling a physical map of this chromosome. Here we report the sequence of two cosmids representing approximately 5% of the polytenized region. Both cosmid clones contain numerous repeated DNA sequences, as identified by cross hybridization with labeled genomic DNA, BLAST searches, and dot matrix analysis, which are positioned between and within the transcribed sequences. The repetitive sequences include three copies of the mobile element Hoppel, one copy of the mobile element HB, and 18 DINE repeats. DINE is a novel, short repeated sequence dispersed throughout both cosmid sequences. One cosmid includes the previously described cubitus interruptus (ci) gene and two new genes: that a gene with a predicted amino acid sequence similar to ribosomal protein S3a which is consistent with the Minute(4)101 locus thought to be in the region, and a novel member of the protein family that includes plexin and met-hepatocyte growth factor receptor. The other cosmid contains only the two short 5'-most exons from the zinc-finger-homolog-2 (zfh-2) gene. This is the first extensive sequence analysis of noncoding DNA from chromosome 4. The distribution of the various repeats suggests its organization is similar to the beta-heterochromatic regions near the base of the major chromosome arms. Such a pattern may account for the diffuse banding of the polytene chromosome 4 and the variegation of many P-element transgenes on the chromosome.

Amino Acid Sequence↗
PubMed2002

Ancient lineages of non-LTR retrotransposons in the primitive eukaryote, Giardia lamblia.

Mobile elements that use reverse transcriptase to make new copies of themselves are found in all major lineages of eukaryotes. The non-long terminal repeat (non-LTR) retrotransposons have been suggested to be the oldest of these eukaryotic elements. Phylogenetic analysis of non-LTR elements suggests that they have predominantly undergone vertical transmission, as opposed to the frequent horizontal transmissions found for other mobile elements. One prediction of this vertical model of inheritance is that the oldest lineages of eukaryotes should exclusively harbor the oldest lineages of non-LTR retrotransposons. Here we characterize the non-LTR retrotransposons present in one of the most primitive eukaryotes, the diplomonad Giardia lamblia. Two families of elements were detected in the WB isolate of G. lamblia currently being used for the genome sequencing project. These elements are clearly distinct from all other previously described non-LTR lineages. Phylogenetic analysis indicates that these Genie elements (for Giardia early non-LTR insertion element) are among the oldest known lineages of non-LTR elements consistent with strict vertical descent. Genie elements encode a single open reading frame with a carboxyl terminal endonuclease domain. Genie 1 is site specific, as seven to eight copies are present in a single tandem array of a 771-bp repeat near the telomere of one chromosome. The function of this repeat is not known. One additional, highly divergent, element within the Genie 1 lineage is not located in this tandem array but is near a second telomere. Four different telomere addition sites could be identified within or near the Genie elements on each of these chromosomes. The second lineage of non-LTR elements, Genie 2, is composed of about 10 degenerate copies. Genie 2 elements do not appear to be site specific in their insertion. An unusual aspect of Genie 2 is that all copies contain inverted repeats up to 172 bp in length.

Animals↗
PubMed1999

Genome system architecture and natural genetic engineering in evolution.

Molecular genetics teaches three lessons relevant to the nature of genetic change during evolution: (1) Genomes are organized as hierarchies of composite systems (multidomain protein-coding sequences; functional loci made up of regulatory, coding, processing, and intervening sequences; and multilocus regulons and replicons) interconnected and organized into specific "system architectures" by repetitive DNA elements. (2) Genetic change often occurs via natural genetic engineering systems (cellular biochemical functions, such as recombination complexes, topoisomerases, and mobile elements, capable of altering DNA sequence information and joining together different genomic components). (3) The activity of natural genetic systems is regulated by cellular control circuits with respect to the timing, activity levels, and specificities of DNA rearrangements (e.g., adaptive mutation, Ty element mobility, and P factor insertions). These three lessons provide plausible molecular explanations for the episodic, multiple, nonrandom DNA rearrangements needed to account for the evolution of novel genomic system architectures and complex multilocus adaptations. This molecular genetic perspective places evolutionary change in the biologically responsive context of cellular biochemistry.

Adaptation, Biological↗
PubMed2006

Applying mobile genetic elements for genome analysis and evolution.

Transposable elements (TEs) are ubiquitous components of all living organisms, and in the course of their coexistence with their respective host genomes, these parasitc DNAs have played important roles in the evolution of complex genetic networks. The interaction between mobile DNAs and their host genomes are quite diverse, ranging from modifications of gene structure and regulation to alterations in general genome architecture. Thus during evolutionary time these elements can be regarded as natural molecular tools in shaping the organization, structure, and function of eukaryotic genes and genomes. Based on their intrinsic properties and features, mobile DNAs are widely applied at present as a technical "toolbox," essential for studying a diverse spectrum of biological questions. In this review, we aim to summarize both the evolutionary impact of TEs on genome evolution and their valuable and diverse methodological applications as molecular tools.

Animals↗
PubMed2004

Mobile genetic elements as natural tools for genome evolution.

Transposable elements (TEs) are ubiquitous components of all living organisms, and in the course of their coexistence with their respective host genomes, these parasitic DNAs have played important roles in the evolution of complex genetic networks. The interaction between mobile DNAs and their host genomes are quite diverse, ranging from modifications of gene structure and regulation to alterations in general genome architecture. Thus over evolutionary time these elements can be regarded as natural molecular tools in shaping the organization, structure, and function of eukaryotic genes and genomes. Based on their intrinsic properties and features, mobile DNAs are widely applied at present as a technical "toolbox," essential for studying a diverse spectrum of biological questions. In this chapter we aim to review both the evolutionary impact of TEs on genome evolution and their valuable and diverse methodological applications as the molecular tools presented in this book.

Alu Elements↗
PubMed1992

Intragenic suppression: Stalker, a retrovirus-like transposable element, can compensate for a deficiency at the cut locus of Drosophila melanogaster.

A number of mutations at the cut locus were induced by non-precise excision of a silent P-element insertion which resulted in deletions at the regulatory region of the locus. Unexpectedly, a reversion of one of these mutations was found, which appears as a result of insertion of Stalker (a retrovirus-like mobile element) near the 1.3 kb deletion. Thus an insertion of a retrovirus-like mobile element can suppress the deficiency at the regulatory region of a gene.

Animals↗
PubMed2004

Gram-positive bacteria are a major reservoir of Class 1 antibiotic resistance integrons in poultry litter.

Reversing the spread of antibiotic multiresistant bacteria is hampered by ignorance of the natural history of resistance genes, the mobile elements carrying them, and the bacterial hosts harboring them. Using traditional cultivation and cultivation-independent molecular techniques, we quantified antibiotic resistance genes and mobile elements called integrons in poultry house litter from commercial poultry farms. Unexpectedly, the major reservoir for Class 1 integrons in poultry litter is not their previously identified hosts, Gram-negative Enterobacteriaceae such as Escherichia coli. Rather, integrons and associated resistance genes abound in several genera of Gram-positive bacteria that constitute >85% of the litter community compared with Enterobacteriaceae that comprise <2% of this ecosystem. This finding warrants reexamination of our assumptions about the persistence and spread of antibiotic resistance genes.

Animals↗
PubMedDate not supplied

[Genome organization and primary structure of the BamHI fragment of highly repetitive DNA from Hordeum vulgare].

The primary structure of three highly repetitive DNA sequences, which are: the most widespread repeats in a number of Hordeum species, represented by 10(5) copies in the genome of Hordeum vulgare, and belong to the BamHI fragment, was studied. The organization of this fragment in the barley genome was described. The computer analysis of the primary structure indicated a great number of inverted repeats (two of them are flanked by short direct repeats) in the BamHI fragment and revealed the short subrepeats of the A, B, C types, five potential RNA-polymerase II sites, one site for RNA polymerase III, open reading frame of 294 b.p. or 210 b.p., sites for polyadenylation and enhancer sequence. The presence of a mobile element (95-568 bp) is very likely in the BamHI fragment, taking into account the location of the mentioned structures along the fragment. This mobile element contains the subrepeats of only A and B types, all sites of transcription initiation, the open reading frame and the inverted repeats on both ends, flanked by the short direct repeats. The transposition mechanisms probably play an essential part in the origin and propagation of the studied BamHI fragment. The presence of potential functional sites in the fragment indicates the possibility of transcription and translation of some highly repetitive Hordeum vulgare DNA.

Blotting, Southern↗