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Repetitive DNA elements characteristic of pathogenic Entamoeba histolytica strains can also be detected after polymerase chain reaction in a cloned nonpathogenic strain.

Strains of Entamoeba histolytica which were isolated from symptomatic patients and which possess a characteristic pathogenic isoenzyme pattern (zymodeme) have extrachromosomal circular DNA molecules containing RNA genes and clusters of tandemly reiterated PvuI elements. The nucleotide sequence of comparable reiterated BamHI elements present in amebae with nonpathogenic zymodemes differs from that found in pathogenic ones. By using the polymerase chain reaction, it was demonstrated that the cloned, nonpathogenic E. histolytica strain SAW 1734R clAR also contains one or few of the tandemly repeated DNA PvuI elements characteristic of the pathogenic amebae. Sequences were detected by hybridization with the P-145 probe after in vitro amplification. Because of technical difficulties, it was impossible to resolve whether single copies of the nonpathogenic BamHI repetitive elements are present in pathogenic amebae. Our findings suggest that in the nonpathogenic amebae, the signal to start amplifying the PvuI-type elements may be induced during the process of elimination of bacterial associates from their growth environment.

Animals↗

The structure of HSAG-1, a middle repetitive genetic element which elicits a leukemia-related cellular surface antigen.

HSAG-1 is a cloned member of a heterogeneous middle repetitive family of genetic elements which is capable of eliciting a leukemia-related surface antigen detected with a monoclonal antibody after DNA transformation of mouse cells. HSAG-1 was originally isolated from a Chinese hamster-human leukemia hybrid cell gene library both by sib-selection for antigen producing activity and by hybridization with labelled human genomic human DNA. We show here that the human labelled site is at the right hand end of the insert, while the antigen-eliciting portion is included in a 1450 bp fragment at the left hand end of the insert. We also present the complete nucleotide sequence of the 3369 bp insert. The sequence contains 12 elements which bear a significant resemblance to accepted consensus sequences for Alu repetitive elements. The right hand end contains adjacent elements with close sequence similarity to portions of the human and hamster type I and type II Alu consensus sequences. All of the other Alu-related elements have diverged relative to the Alu consensus sequences by additions, long deletions and substitutions. The left hand portion of the insert which has the antigen-producing activity contains four of these diverged elements representing a relatively high proportion (26%) of the nucleotide sequence. The sequence is thus consistent with our previous observations of a repetitive family with biological function.

Animals↗

MaskerAid: a performance enhancement to RepeatMasker.

UNLABELLED: Identifying and masking repetitive elements is usually the first step when analyzing vertebrate genomic sequence. Current repeat identification software is sensitive but slow, creating a costly bottleneck in large-scale analyses. We have developed MaskerAid, a software enhancement to RepeatMasker that increased the speed of masking more than 30-fold at the most sensitive setting. AVAILABILITY: On request from the authors (see http://sapiens.wustl.edu/MaskerAid). CONTACT: maskeraid@watson.wustl.edu

Animals↗

DNA nucleotide sequence analysis of the PvuII DNA fragment L of the genome of insect iridescent virus type 6 reveals a complex cluster of multiple tandem, overlapping, and interdigitated repetitive DNA elements.

The DNA nucleotide sequence of the PvuII DNA fragment L (0.920 to 0.944 map units (m.u.] of the genome (209 kbp) of insect iridescent virus type 6 was determined. The size of this DNA fragment was 5064 bp with a base composition of 39.79% G + C and 60.21% A + T. The DNA sequence contained many perfect direct repeats of sizes up to 145 bp. In addition to these repetitions, a cluster of four imperfect repetitive DNA elements (R1 to R4) with a complex structural arrangement was detected. R1, R2, and R3 existed in duplicate (two boxes (B] between nucleotide positions 271 and 3466) and their size were as follows: R1-B1/B2 (567/568 bp), R2-B1/B2 (917/931 bp), and R3-B1/B2 (92/88 bp). The R4 repetitive element was found in 12 boxes (between bases 1301 and 4417), which were interrupted at nucleotide positions 1883 to 2236 and 3341 to 3587. These interruptions define three segments (S) harboring boxes B1 to B3 (S1), B4 to B8 (S2), and B9 to B12 (S3). The size of the individual boxes was found to be 239, 233, 107, 244, 222, 242, 242, 148, 240, 242, 242, and 102 bp for R4-B1 to B12, respectively. Five open reading frames (ORFs of 118 to 333 amino acid (AA) residues) were detected. The analysis of the amino acid sequences of the largest ORF revealed that the deduced amino acid sequence of the putative gene product contained two repetitions TR1 (three domains of 50 AA) and TR2 (two domains of 74 AA). Sequences of 43 amino acid residues of ORF 5 (160 to 202 AA) were homologous within the majority of ORFs. A consensus sequence-MANL(X)6 IGSSST(X)6 L(X)1 LGS(X)1 LQISG(X)2 L(X)1 VN- was found in all five ORFs. Although classical canonical and noncanonical transcriptional start signals were detectable, polyadenylation signals were not observed.

Amino Acid Sequence↗

A cytomegalovirus DNA sequence containing tracts of tandemly repeated CA dinucleotides hybridizes to highly repetitive dispersed elements in mammalian cell genomes.

A single 880-base-pair region within the genome of simian cytomegalovirus strain Colburn contains sequences that hybridize intensely with both human and mouse total genome DNA probes. This sequence was also found in a second simian cytomegalovirus isolate and was retained in both plaque-purified virus subclones and in plasmid DNA clones containing the SalI P fragment. Cleaved genomic DNAs from several mammalian species all exhibited strong dispersed hybridization with the SalI-P probes, and over 70% of the lambda clones in a mouse genomic library plus several selected clones containing globin, 45S rDNA, or 5S rDNA genes all formed hybrids with SalI-P. The appropriate region of cytomegalovirus SalI-P contains relatively A + T-rich unique sequences interrupted by three stretches of the simple alternating dinucleotides, (CA)15, (CA)22, and (CA)21, which we show to be responsible for most of the cell-virus homology. We conclude that discrete, tandemly repeated (CA) dinucleotide tracts capable of forming left-handed Z-DNA helices punctuate mammalian genomes at greater than 10(5) copies per cell and that three adjacent copies of what appear to be a family of interspersed repetitive elements containing these (CA)n stretches are carried in the genomes of simian cytomegaloviruses.

Animals↗

On the roles of repetitive DNA elements in the context of a unified genomic-epigenetic system.

Repetitive DNA sequences comprise a substantial portion of most eukaryotic and some prokaryotic chromosomes. Despite nearly forty years of research, the functions of various sequence families as a whole and their monomer units remain largely unknown. The inability to map specific functional roles onto many repetitive DNA elements (REs), coupled with the taxon-specificity of sequence families, have led many to speculate that these genomic components are "selfish" replicators generating genomic "junk." The purpose of this paper is to critically examine the selfishness, evolutionary effects, and functionality of REs. First, a brief overview of the range of ideas pertaining to RE function is presented. Second, the argument is presented that the selfish DNA "hypothesis" is actually a narrative scheme, that it serves to protect neo-Darwinian assumptions from criticism, and that this story is untestable and therefore not a hypothesis. Third, attempts to synthesize the selfish DNA concept with complex systems models of the genome and RE functionality are critiqued. Fourth, the supposed connection between RE-induced mutations and macroevolutionary events are stated to be at variance with empirical evidence and theoretical considerations. Hypotheses that base phylogenetic transitions in repetitive sequence changes thus remain speculative. Fifth and finally, the case is made for viewing REs as integrally functional components of chromosomes, genomes, and cells. It is argued throughout that a new conceptual framework is needed for understanding the roles of repetitive DNA in genomic/epigenetic systems, and that neo-Darwinian "narratives" have been the primary obstacle to elucidating the effects of these enigmatic components of chromosomes.

Animals↗

Structure and function of repetitive sequence elements associated with a highly polymorphic domain of the Neisseria meningitidis PilQ protein.

Secretins are a large family of proteins associated with membrane translocation of macromolecular complexes, and a subset of this family, termed PilQ proteins, is required for type IV pilus biogenesis. We analysed the status of PlIQ expression in Neisseria meningitidis (Mc) and found that PlIQ mutants were non-piliated and deficient in the expression of pilus-associated phenotypes. Sequence analysis of the 5' portion of the pilQ ORF of the serogroup B Mc strain 44/76 showed the presence of seven copies of a repetitive sequence element, in contrast to the situation in N. gonorrhoeae (Gc) strains, which carry either two or three copies of the repeat. The derived amino acid sequence of the consensus nucleotide repeat was an octapeptide PAKQQAAA, designated as the small basic repeat (SBR). This gene segment was studied in more detail in a collection of 52 Mc strains of diverse origin by screening for variability in the size of the PCR-generated DNA fragments spanning the SBRs. These strains were found to harbour from four to seven copies of the repetitive element. No association between the number of copies and the serogroup, geographic origin or multilocus genotype of the strains was evident. The presence of polymorphic repeat elements in Mc PilQ is unprecedented within the secretin family. To address the potential function of the repeat containing domain, Mc strains were constructed so as to express chimeric PilQ molecules in which the number of SBR repeats was increased or in which the repeat containing domain was replaced in toto by the corresponding region of the Pseudomonas aeruginosa (Pa) PilQ protein. Although the strain expressing PilQ with an increased number of SBRs was identical to the parent strain in pilus phenotypes, a strain expressing PilQ with the equivalent Pa domain had an eightfold reduction in pilus expression level. The findings suggest that the repeat containing domain of PilQ influences Mc pilus expression quantitatively but not qualitatively.

Amino Acid Sequence↗

Invasion of the human albumin-alpha-fetoprotein gene family by Alu, Kpn, and two novel repetitive DNA elements.

The human albumin-alpha-fetoprotein genomic domain contains 13 repetitive DNA elements randomly distributed throughout the symmetrical structures of these genes. These repeated sequences are located at different sites within the two genes. The human albumin gene contains five Alu elements within four of its 14 intervening sequences. Two of these repeats are located in intron 2, and the remaining three are located in introns 7, 8, and 11. The human alpha-fetoprotein gene contains three of these Alu elements, one in intron 4 and the remaining two in the 3'-untranslated region. In addition, the human alpha-fetoprotein gene contains a Kpn repeat and two classes of novel repeats that are absent from the human albumin gene. Six of the Alu elements within the two genes are bound by short direct repeats that harbor five base substitutions in 120 possible positions (60 bp times 2 termini). The absence of Alu repeats from analogous positions in rodents indicates that these repeats invaded the albumin-alpha-fetoprotein domain less than 85 Myr ago (the time of mammalian radiation). Furthermore, considering the conservation of terminal repeats flanking the Alu sequences of the albumin-alpha-fetoprotein domain (0.042 changes per site), we submit that the average time of Alu insertion into this gene family could have been as recently as 15-30 Myr ago.

Base Sequence↗

Molecular evidence for a relationship between LINE-1 elements and X chromosome inactivation: the Lyon repeat hypothesis.

X inactivation is a chromosome-specific form of genetic regulation in which thousands of genes on one homologue become silenced early in female embryogenesis. Although many aspects of X inactivation are now understood, the spread of the X inactivation signal along the entire length of the chromosome remains enigmatic. Extending the Gartler-Riggs model [Gartler, S. M. & Riggs, A. D. (1983) Annu. Rev. Genet. 17, 155-190], Lyon recently proposed [Lyon, M. F. (1998) Cytogenet. Cell Genet. 80, 133-137] that a nonrandom organization of long interspersed element (LINE) repetitive sequences on the X chromosome might be responsible for its facultative heterochromatization. In this paper, we present data indicating that the LINE-1 (L1) composition of the human X chromosome is fundamentally distinct from that of human autosomes. The X chromosome is enriched 2-fold for L1 repetitive elements, with the greatest enrichment observed for a restricted subset of LINE-1 elements that were active <100 million years ago. Regional analysis of the X chromosome reveals that the most significant clustering of these elements is in Xq13-Xq21 (the center of X inactivation). Genomic segments harboring genes that escape inactivation are significantly reduced in L1 content compared with X chromosome segments containing genes subject to X inactivation, providing further support for the association between X inactivation and L1 content. These nonrandom properties of L1 distribution on the X chromosome provide strong evidence that L1 elements may serve as DNA signals to propagate X inactivation along the chromosome.

Base Pairing↗

Transposable elements and vertebrate protein diversity.

Interspersed repetitive sequences are major components of eukaryotic genomes. Repetitive elements comprise about 50% of the mammalian genome. They interact with the whole genome and influence its evolution. Repetitive elements may serve as recombination hot spots or acquire specific cellular functions such as RNA transcription control or become part of protein coding regions. The latter is a subject of presented analysis. We searched all currently available vertebrate protein sequences, including human proteome complement for the presence of transposable elements. It appears that insertion of TE-cassettes into open reading frames is a general phenomena. They can be found in all vertebrate lineages and originate in all types of transposable elements. It seems that genomes use those cassettes as 'ready to use' motifs in their evolutionary experiments. Most of TE-cassettes are used to create alternative forms of a message and usually the other form, without TE-cassette, is expressed in a cell. Tables listing vertebrate messages with TE-cassettes are available at http://warta.bio.psu.edu/ScrapYard/.

Animals↗

Genome structure of spirochetes.

The genome structures of several pathogenic spirochetes have recently been determined. The genomes of Borrelia species consist of a linear chromosome of approximately one million base pairs (Mb) and various linear and circular plasmids. Analysis of restriction fragment length polymorphisms and 16S ribosomal RNA sequence data indicate the division of Borrelia burgdorferi into at least three distinct genetic groups. Leptospira interrogans has a circular chromosome 5 Mb in size and a 0.35 Mb extrachromosomal element. Repetitive sequence elements similar to insertion sequences have been identified in the Leptospira interrogans genome. The chromosome of Treponema pallidum subsp. pallidum is circular and has a size of approximately one Mb. Genetic studies conducted to date indicate that B. burgdorferi and L. interrogans have a high degree of genetic diversity, whereas remarkably few genetic differences have been observed among the pathogenic Treponema. Knowledge of the genomic structure of these organisms will serve as a basis for future genetic studies.

Borrelia burgdorferi Group↗

Current understanding of the genetic diversity of Mycobacterium avium subsp. paratuberculosis.

Mycobacterium avium subsp. paratuberculosis (MAP) is the etiological agent of Johne's disease (or paratuberculosis). Paratuberculosis is a chronic gastroenteritis mainly affecting cattle, sheep and other ruminants. MAP is also of concern due to the heretofore unresolved issue of its possible role in Crohn's disease in humans. We present here a review of MAP (i) mobile genetic elements; (ii) repetitive elements; (iii) single nucleotide polymorphisms; and (iv) whole-genome comparisons to study the molecular epidemiology of MAP. A summary of the findings to date is presented, and the discriminatory power, advantage and disadvantages of each of the methods are compared and discussed.

Animals↗

Isolation and initial characterization of the alcohol dehydrogenase gene from Drosophila affinidisjuncta.

Recombinant bacteriophages containing the alcohol dehydrogenase (ADH) gene from Drosophila affinidisjuncta have been isolated by virtue of their cross-hybridization to the previously cloned ADH gene from D. melanogaster. Within the 17 kilobases of cloned DNA represented in the phage genomes, the sequences hybridizing to the D. melanogaster ADH gene lie roughly in the center. The only region of detectable hybridization to cDNA made from templates of D. affinidisjuncta larval poly(A)-containing RNA maps to the same portion of the cloned DNA. Verification that the phages carry the ADH structural gene was obtained by hybrid-selecting ADH mRNA, translating it in vitro, and immunoprecipitating the resulting ADH polypeptide. Analysis of genomic DNA suggests that the ADH gene and most flanking sequences are present only once in the haploid genome. However, 3' to the ADH gene, two separable repetitive elements are found. Both repetitive elements are probably small and poorly conserved in the genome, and neither interferes with localization of the ADH gene, by in situ hybridization, to a position near the base of the third chromosome. Analysis of ADH transcripts demonstrates that there are at least four RNAs produced by the ADH gene. Two size classes of RNA are seen at each stage of development. In addition, ADH transcripts from larvae and adults differ from one another in a reproducible manner.

Alcohol Dehydrogenase↗

Structural analysis of repetitive sequence elements transcribed in early development of Xenopus laevis.

A cDNA library prepared from mRNA of Xenopus laevis embryos was screened with a genomic DNA fragment containing various transcribed repetitive sequence elements. Comparison of the nucleotide sequence of two isolated cDNAs and their genomic relatives allows one to define two transcribed repetitive sequence elements. One of them belongs to a highly reiterated family and consists of a tandem array of homologous subunits of 77-80 bp. The other is reiterated approximately 2200 times, has a size of 260 bp and displays a conserved region of 135 bp. The data are consistent with the presence of repetitive sequence transcripts in the 3' part of mRNA molecules.

Animals↗

Use of a mammalian interspersed repetitive (MIR) element in the coding and processing sequences of mammalian genes.

The mammalian interspersed repetitive (MIR) element was amplified in mammals 130 million years ago. The MIR element is at least 260 bp in length and is found in approximately 105 copies in the mammalian genome. We analyzed copies of the MIR element in the DNA of various mammals to determine its relationship to the structure and function of genes, in an attempt to identify specific uses of the MIR element within the mammalian genome. We found that alternative splicing within the acetylcholine receptor gene in humans takes place within the MIR element and results in the incorporation of part of the MIR element into the coding sequence of this gene. Furthermore, the polyadenylation signal (AATAAA) at the 3' end of four different mammalian genes is derived from the MIR element. These uses of the MIR element suggest that other regulatory sequences found within the mammalian genome originated from ancient transposable elements, many of which may no longer be recognizable.

Alternative Splicing↗

Exclusion of repetitive DNA elements from gnathostome Hox clusters.

Despite their homology and analogous function, the Hox gene clusters of vertebrates and invertebrates are subject to different constraints on their structural organization. This is demonstrated by a drastically different distribution of repetitive DNA elements in the Hox cluster regions. While gnathostomes have a strong tendency to exclude repetitive DNA elements from the inside of their Hox clusters, no such trend can be detected in the Hox gene clusters of protostomes. Repeats "invade" the gnathostome Hox clusters from the 5' and 3' ends while the core of the clusters remains virtually free of repetitive DNA. This invasion appears to be correlated with relaxed constraints associated with gene loss after cluster duplications.

Animals↗

Densities, length proportions, and other distributional features of repetitive sequences in the human genome estimated from 430 megabases of genomic sequence.

The densities of repetitive elements in the human genome were calculated in each GC content class using non-overlapping windows of 50kb. The density of Alu is two to three times higher in GC-rich regions than in AT-rich regions, while the opposite is true for LINE1. In contrast, LINE2 and other elements, such as DNA transposons, are more uniformly distributed in the genome. The number of Alus in the human genome was estimated to be 1.4 million, higher than previous estimates. About 40% of the autosomes and approximately 51% of the X and Y chromosomes are occupied by repetitive elements. In total, the human genome is estimated to contain more than 4 million repetitive elements. The GC contents (%) of repetitive elements and their flanking regions were also calculated. The GC contents of almost all kinds of repeats are positively correlated with the window GC contents, suggesting that a repetitive sequence is subject to the same mutation pressure as its surrounding regions, so it tends to have the same GC content as its surrounding regions. This observation supports the regional mutation hypothesis. The only two exceptions are AluYa and AluYb8, the two youngest Alu subfamilies. The GC content of AluYb8 is negatively correlated with that of its surrounding regions, while AluYa shows no correlation, suggesting different insertion patterns for these two young Alu subfamilies. This suggestion was supported by the fact that the average genetic distance between members of AluYb8 in each GC window class is positively correlated with the GC content of the window, but no correlation was found for AluYa. AluYa is more frequent in Y chromosome than in other chromosomes; the same is true for LTR retroviruses. This pattern might be correlated with the evolutionary history of Y chromosome.

Alu Elements↗

Genomic scrap yard: how genomes utilize all that junk.

Interspersed repetitive sequences are major components of eukaryotic genomes. Repetitive elements comprise over 50% of the mammalian genome. Because the specific function of these elements remains to be defined and because of their unusual 'behaviour' in the genome, they are often quoted as a selfish or junk DNA. Our view of the entire phenomenon of repetitive elements has to now be revised in the light of data on their biology and evolution, especially in the light of what we know about the retroposons. I would like to argue that even if we cannot define the specific function of these elements, we still can show that they are not useless pieces of the genomes. The repetitive elements interact with the surrounding sequences and nearby genes. They may serve as recombination hot spots or acquire specific cellular functions such as RNA transcription control or even become part of protein coding regions. Finally, they provide very efficient mechanism for genomic shuffling. As such, repetitive elements should be called genomic scrap yard rather than junk DNA. Tables listing examples of recruited (exapted) transposable elements are available at http://www.ncbi.nlm.gov/Makalowski/ScrapYard/

Animals↗