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PubMed1985

Inverted terminal repeats and terminal proteins of the genomes of pneumococcal phages.

The nucleotide (nt) sequence at the ends of the genomes of the Streptococcus pneumoniae phages Cp-5 and Cp-7 has been determined and compared with the corresponding sequence of phage Cp-1. The genomes of phages Cp-5 and Cp-7 have inverted terminal repeats (ITRs) 343 and 347 bp long, respectively. In Cp-1 DNA the ITR is 236 bp long and the following 116 bp are 93% homologous. Some regions within the ITRs are conserved in the three genomes although the complete sequence of the ITRs is no more conserved than the rest of their genomes. The chromatographic behavior of their tryptic peptides suggests that the terminal proteins (TPs) of at least two of the phages are similar and that the TPs of the three pneumococcal phages differ markedly from that of the Bacillus subtilis phage psi 29.

Bacteriophages↗
PubMed1987

Multiple reiteration of a 40-bp nucleotide sequence in the inverted terminal repeat of the genome of a canine adenovirus.

The DNA of a vaccine strain of canine adenovirus type 1 [ICHV vaccine; Connaught Laboratories, Ltd.; CAV-1(CLL)] has been cloned in plasmid pAT153 in the form of subgenomic BamHI digestion fragments. Analysis of the nucleotide sequences of cloned terminal fragments has revealed an inverted terminal repeat (ITR) with a minimum length of 198 nucleotides, including a tandem reiteration of the 40-bp nucleotide sequence from positions 14 to 53. The ITRs had the 5'-CATCATCAAT ... sequence typical of adenoviruses and the highly conserved sequence ATAATATAC (nucleotides 9-17) of human strains. Additionally, one BamHI A clone (left terminus) contained three sequential copies of the 40-bp sequence, and two BamHI C clones (right terminus) contained at least seven. These did not appear to be artifacts of cloning, since evidence was obtained that the multiple reiterations also occurred in DNA isolated from intact virus. By analogy with human adenoviruses, the repetitive sequence in the CAV-1(CLL) genome encompasses the entire nuclear factor I (NFI) binding site of the origin of DNA replication. Additionally, the 40-bp nucleotide sequence was found to contain the sequence AGG(N)4GCCTAA (nucleotides 27-39), which closely resembles the concensus sequence of the human adenovirus NFI binding site [TGG(N)6-7GCCAA; nucleotides 25-38]. It appears, therefore, that the Connaught CAV-1 vaccine contains reiterated copies of an essential part of the adenoviral origin of DNA replication. A mechanism is proposed for the generation of multiple reiterations of sequences in the right ITR, given an initial single tandem repeat in the left ITR.

Adenoviridae↗
PubMed1981

Terminal proteins and short inverted terminal repeats of the small Bacillus bacteriophage genomes.

The genome of Bacillus phage phi 29 contains covalently linked protein at both ends. These DNA terminal proteins are essential for phi 29 DNA replication. We have isolated phi 29 terminal protein from each end separately and compared their two-dimensional peptide maps. Our results showed the two proteins to be identical. The DNAs of four phages examined (phi 15, Nf, M2Y, and GA-1) also contain protein at both ends of the DNA molecules. The chymotryptic peptide maps of these DNA terminal proteins have been compared with the map of the phi 29 terminal protein. Despite the similarities in molecular size, peptide maps of the terminal proteins show clear differences among the unrelated phages. These results are consistent with the idea that the terminal proteins are encoded by viral DNA rather than by the host chromosome. We have also determined the nucleotide sequences of the termini of four phage DNAs and compared them with the sequence of phi 29 DNA. The sequence data indicate that all of these phages DNA contain short inverted terminal repeats: 5'A-A-A-G-T-A for phi 29 and phi 15, 5' A-A-A-G-T-A-A-G for Nf and M2Y, and 5' A-A-A-T-A-G-A for GA-1.

Bacteriophages↗
PubMed1990

Replication of an adenovirus type 34 mutant DNA containing tandem reiterations of the inverted terminal repeat.

A mutant of human adenovirus type 34 (Ad34) has been isolated which contains DNA molecules with tandem reiterations of from two to eight copies of a 131-bp sequence within the right-sided inverted terminal repetition. Terminal heterogeneity was not eliminated by repeated plaque purifications indicating that the population of DNA molecules with various numbers of reiterations could rapidly evolve from the DNA of a single virus particle. These enlarged DNA molecules were capable of replication both in vivo and in vitro. The nucleotide sequence of the mutant Ad34 inverted terminal repetitions contained most of the essential features of the Ad origin of DNA replication. These features include the ATAATATACC sequence which is present between the highly conserved bases 9-18 in all human adenoviruses, as well as the consensus sequences for the binding of nuclear factor I and nuclear factor III. However, the reiterated sequences lacked a dG appropriately placed on the template strand to serve as a potential site for internal initiation. It appears that the rapid amplification of two to eight copies of the reiterated terminal sequences does not arise from internal initiation during replication but probably from homologous recombination.

Adenoviruses, Human↗
PubMed1987

Genetic analysis of the interaction of the insertion sequence IS903 transposase with its terminal inverted repeats.

The insertion sequence IS903 has perfect, 18-base-pair terminal repeats that are the presumed binding sites of its transposase. We have isolated mutations throughout this inverted repeat and analyzed their effect on transposition. We show that every position in the inverted repeat (with the possible exception of position 4) is important for efficient transposition. Furthermore, various substitutions at a single position can have a wide range of effects. Analysis of these hierarchical effects suggests that transposase contacts the minor groove in the region from position 13 to position 16 but makes major groove (or more complex) interactions with the outer portion of the inverted repeat. Our data indicate that the transposase exhibits relaxed specificity for the "second" end of a transposed segment; the defect in transposition of virtually all mutant inverted repeats can be rescued by a wild-type end. However, this rescue exhibits a pronounced position effect; in most cases, it is efficient only when the wild-type end is close to the 3' end of the transposase gene. This confirms the cis-acting nature of the transposase protein and suggests the initial transposase-inverted repeat interaction is the rate-limiting step in transposition. From the behavior of transposons with one mutant and one wild-type end, we infer that the inverted repeat contains two functional domains--one for initial complex formation with transposase and the other for effective completion of transpositional recombination. To support this hypothesis we show that an end with a mutation in one domain can significantly rescue an end with a mutation in the other domain.

Binding Sites↗
PubMed1984

Dictyostelium transposable element DIRS-1 has 350-base-pair inverted terminal repeats that contain a heat shock promoter.

DIRS-1 is a 4.7-kilobase-pair repetitive and apparently transposable Dictyostelium genetic element that is transcribed during differentiation or after heat shock. The terminal regions of DIRS-1 are inverted repeats of 330 base pairs. The repeats are highly conserved both within a given element as well as between different members of the family (less than 10% divergence). At the distal end of all left repeats is a 32-nucleotide sequence composed almost entirely of A and T residues. In addition to this 32-base A + T sequence, the distal region of all right repeats is extended by a 28-base-pair A + T-rich sequence that is identical in all copies. The sequences flanking each DIRS-1 sequence are completely dissimilar, and there appears to be no duplication of the genomic DNA sequence at the presumed point of DIRS-1 insertion. The terminal repeats can also be found interspersed in the genome independently of the complete element. In addition, the terminal repeats carry a 15-nucleotide sequence that greatly resembles the Drosophila consensus heat shock promoter and may be involved in the transcriptional induction of the DIRS-1 sequences.

Base Sequence↗
PubMed1991

Stability of molluscum contagiosum virus DNA among 184 patient isolates: evidence for variability of sequences in the terminal inverted repeats.

The stability of the Molluscum contagiosum virus Type 1 genome (188 kbp) was studied in 184 DNA isolates from 131 patients. Variability of up to 1.5 kbp at both ends of the genome symmetrically was observed using restriction analysis of the DNA isolates and by Southern Blot experiments using cloned and labeled HindIII terminal DNA fragments of MCV-1 prototype DNA. The variable sequences were mainly confined to the terminal fragments and parts of the MCV-1 terminal repeats. Labeled probes did not detect terminal sequences of MCV Type 2 under the applied stringency. A less marked instability of the central MCV-1 BamHI DNA fragment F was observed within the genome coordinates 0.431 to 0.454 mu. Reiteration of tandem repeats similar to those described for vaccinia virus might explain the variability of the terminal sequences and might be involved in viral replication.

Blotting, Southern↗
PubMed1990

The N-terminal domain of the insertion sequence 30 transposase interacts specifically with the terminal inverted repeats of the element.

The gene for the insertion sequence (IS) 30 transposase is placed under the control of the tac promoter, and large quantities of transposase are expressed upon induction. The resulting protein precipitates inside the Escherichia coli cells in the form of inclusion bodies which, upon cell lysis, cannot be dissolved under nondenaturing conditions. In contrast, the N-terminal third of the transposase, a 17-kDa protein produced by a truncated gene, can be purified and is able to interact site specifically with the ends of the IS30 element. In DNase I footprint experiments, regions of 26 nucleotides on one DNA strand and 19 nucleotides on the other strand at either end of the element are protected from nuclease digestion. It is concluded that a functional DNA-binding domain can be formed by expression of only one-third of the complete IS30 transposase. Sequence comparison shows a homology of the IS30 ends to the ends of IS4351 and to the L1 end of bacteriophage Mu.

Amino Acid Sequence↗
PubMed1986

Characterization of an adenovirus type 5 mutant carrying embedded inverted terminal repeats.

During construction of an adenovirus type 5 (Ad5) deletion mutant, dlE1,3, lacking E1 and E3 sequences, we isolated a variant, dlE1,3-1, which had a direct repeat of viral DNA terminal sequences attached to the left end of the genome. Analysis of this variant with restriction enzymes and by hybridization of Southern blots with specific probes indicated that the extra terminal segment contained the left 2.6% (920 bp) of Ad5 joined to 352 bp of pBR322 which in turn was linked to the left end (minus 21 bp) of dlE1,3. During replication of dlE1,3-1 the extra terminal segment was found to transfer to the right end of the genome resulting in a second variant, dlE1,3-2, with duplicated terminal sequences at both ends of the viral genome. DlE1,3-2 in turn was shown to revert back to dlE1,3-1 at high frequency. Although evidence was obtained indicating that the extra segment could be lost from the left end, spontaneous mutants which had lost direct repeats from both ends were never isolated. It was, however, possible to remove the extra terminal repeat of dlE1,3-1 by cleavage with a restriction enzyme and to isolate dlE1,3 containing wt termini. The rearrangements occurring during replication of dlE1,3-1 and dlE1,3-2 may be consequences of the mode of replication of Ad5 DNA and bear some resemblance to segment inversion in herpesviruses.

Adenoviruses, Human↗
PubMed1986

Terminal inverted repeats of prokaryotic transposable element IS186 which can generate duplications of variable length at an identical target sequence.

The insertion element IS186, which resides in the chromosome of Escherichia coli K-12, is 1338 bp long. Its termini represent 23-bp perfectly inverted repeats, but a variant carries a mismatch at position 23. IS186 transposes preferentially into G + C-rich sequences and generates target duplications of variable length, even at the same integration site.

Base Composition↗
PubMed1978

Inverted terminal repeats in rabbit poxvirus and vaccinia virus DNA.

In both rabbit poxvirus and vaccinia virus DNA have demonstrated an identical distribution of eight HinfI. The length of the terminal repeats was found to be 3.4 to 3.6 megadaltons (Mdaltons) for rabbit poxvirus DNA and 7.4 to 8.0 Mdaltons for vaccinia virus DNA. Maps of the HinfI restriction sites within isolated EcoRI end fragments of rabbit poxvirus and vaccinia virus DNA PHAVE DEMONSTRATED AN IDENTICAL DISTRIBUTION OF EIGHT HinfI sites in an internal part (approximately 2 Mdaltons) of the EcoRI end fragments of the two genomes.

Animals↗
PubMed1984

Nucleotide sequence of the maize transposable element Mul.

A cloned DNA fragment from the maize allele Adhl-S3034 contains all of Mul, an insertion element involved in Robertson's Mutator activity. The element is 1367 base pairs (bp) long and is flanked by nine bp direct repeats of insertion site DNA. It has inverted terminal repeats of 215 and 213 bp showing 95% homology. Within the element are two direct repeats of 104 bp showing 96% homology. Four open reading frames (ORFs) were found, two in each DNA strand. Mul can be divided into two halves, each containing one terminal inverted repeat, an internal direct repeat, and two overlapping ORFs. The GC content of each half is high (70%), while that of a central 60 base portion of the element is low (26%). The central region contains the only sequence resembling the TAATA Goldberg and Hogness eukaryotic promoter signal. Multiple copies of DNA sequences related to Mul found in Mutator maize plants are generally similar in organization to the cloned element. A larger version containing a discrete 300 to 400 base pair insertion was found in some Mutator lines.

Base Sequence↗
PubMed1989

Composite transposable elements in the Xenopus laevis genome.

Members of two related families of transposable elements, Tx1 and Tx2, were isolated from the genome of Xenopus laevis and characterized. In both families, two versions of the elements were found. The smaller version in each family (Tx1d and Tx2d) consisted largely of two types of 400-base-pair tandem internal repeats. These elements had discrete ends and short inverted terminal repeats characteristic of mobile DNAs that are presumed to move via DNA intermediates, e.g., Drosophila P and maize Ac elements. The longer versions (Tx1c and Tx2c) differed from Tx1d and Tx2d by the presence of a 6.9-kilobase-pair internal segment that included two long open reading frames (ORFs). ORF1 had one cysteine-plus-histidine-rich sequence of the type found in retroviral gag proteins. ORF2 showed more substantial homology to retroviral pol genes and particularly to the analogs of pol found in a subclass of mobile DNAs that are supposed retrotransposons, such as mammalian long interspersed repetitive sequences, Drosophila I factors, silkworm R1 elements, and trypanosome Ingi elements. Thus, the Tx1 elements present a paradox by exhibiting features of two classes of mobile DNAs that are thought to have very different modes of transposition. Two possible resolutions are considered: (i) the composite versions are actually made up of two independent elements, one of the retrotransposon class, which has a high degree of specificity for insertion into a target within the other, P-like element; and (ii) the composite elements are intact, autonomous mobile DNAs, in which the pol-like gene product collaborates with the terminal inverted repeats to cause transposition of the entire unit.

Amino Acid Sequence↗
PubMed1986

Repetitive DNA and chromosome evolution in plants.

Most higher plant genomes contain a high proportion of repeated sequences. Thus repetitive DNA is a major contributor to plant chromosome structure. The variation in total DNA content between species is due mostly to variation in repeated DNA content. Some repeats of the same family are arranged in tandem arrays, at the sites of heterochromatin. Examples from the Secale genus are described. Arrays of the same sequence are often present at many chromosomal sites. Heterochromatin often contains arrays of several unrelated sequences. The evolution of such arrays in populations is discussed. Other repeats are dispersed at many locations in the chromosomes. Many are likely to be or have evolved from transposable elements. The structures of some plant transposable elements, in particular the sequences of the terminal inverted repeats, are described. Some elements in soybean, antirrhinum and maize have the same inverted terminal repeat sequences. Other elements of maize and wheat share terminal homology with elements from yeast, Drosophila, man and mouse. The evolution of transposable elements in plant populations is discussed. The amplification, deletion and transposition of different repeated DNA sequences and the spread of the mutations in populations produces a turnover of repetitive DNA during evolution. This turnover process and the molecular mechanisms involved are discussed and shown to be responsible for divergence of chromosome structure between species. Turnover of repeated genes also occurs. The molecular processes affecting repeats imply that the older a repetitive DNA family the more likely it is to exist in different forms and in many locations within a species. Examples to support this hypothesis are provided from the Secale genus.

Animals↗
PubMed1988

Identification and purification of a Drosophila protein that binds to the terminal 31-base-pair inverted repeats of the P transposable element.

We have used DNase I footprinting and partially fractionated nuclear extracts from Drosophila Kc tissue culture cells to identify DNA-binding proteins that interact with the terminal repeats of P transposable elements. We have identified a binding activity that interacts specifically with a region of the 31-base-pair terminal inverted repeats that is directly adjacent to the duplication of target site DNA. Binding occurs to both the 5' and 3' inverted terminal repeats irrespective of the sequence of the duplicated target DNA. UV photochemical crosslinking studies suggest that the binding activity resides in a polypeptide of 65-70 kDa. Biochemical fractionation and oligonucleotide affinity chromatography have been used to purify the binding activity to near homogeneity and identify a polypeptide of 66 kDa in the highly purified preparations. The site to which binding occurs is included in a region absolutely required for P element transposition, suggesting that this binding protein may be a cellular factor involved in P element transposition.

Animals↗
PubMed1990

Tn3 transposition immunity is conferred by the transposase-binding domain in the terminal inverted-repeat sequence of Tn3.

A series of mutant terminal inverted repeats (IRs), having 2 bp substitutions at various sites within the 38-bp IR sequence of the ampicillin-resistance transposon Tn3, were tested for transposition immunity to Tn3. Mutations within region 1-10 in the IR did not affect transposition immunity, while mutations within region 13-38 inactivated the immunity function. These two regions corresponded to domain A which was not bound specifically by Tn3 transposase and to domain B which was bound by the transposase, respectively. This indicates that specific binding of transposase to domain B within the IR sequence is responsible for transposition immunity.

Ampicillin Resistance↗