IS870 requires a 5'-CTAG-3' target sequence to generate the stop codon for its large ORF1

Abstract

The TB regions of the Agrobacterium vitis octopine/cucumopine Ti plasmids constitute a family of related structures. All contain a bacterial insertion element downstream of the TB-iaaM gene, IS870.1. Whereas 43 isolates with octopine/cucumopine Ti plasmids carry only one IS870 copy, strain Ag57 carries a second copy (IS870.2) 3.9 kb to the right of IS870.1 and part of the same TB region. Two other octopine/cucumopine strains carry an IS870 copy on their chromosome (IS870.3). A study of the unmodified insertion sites of IS870.2 and IS870.3, cloned from closely related strains, enabled us to delimit the IS870 elements. IS870 has a size of 1,152 bp and is terminated by inverted repeats. It contains a large open reading frame without a stop codon. However, a stop codon is generated by insertion into the target sequence 5'-CTAG-3'. IS870 is related to five other insertion sequence elements. For two of these, the stop codon of the largest open reading frame is also created by insertion into a CTAG target site.Bacterial insertion sequences (IS) play an important role in bacterial evolution (32). Their transposition into new locations may cause mutations by gene disruption but can also create potentially useful variability within a population. We are interested in the role of IS elements in the evolution of Agrobacterium Ti plasmids. Part of the Ti plasmid (its T-DNA or T region) is transferred to the plant cell during tumor induction. Expression of T-DNA genes then leads to tumor formation (for a review, see reference 42). The various octopine/cucumopine (o/c) Ti plasmids found in Agrobacterium vitis (20) isolates code for tumor induction on grapevine. They are particularly suited for evolutionary studies, since they are closely related to each other but nevertheless show considerable variation (for a review, see reference 21). All o/c plasmids harbor two independent T regions: TA and TB. The o/c plasmids have been divided into five different types on the basis of their TA region structure. These structures are derived from an ancestor TA region by insertion of different IS elements (IS866, IS868, and IS869) and by deletions caused by insertion of and recombination between different copies of the same IS element (IS868 and IS867). Two large o/c Ti plasmid groups can be distinguished: the "small TA" group (with its representatives pTiAB3 and pTiAg57) and the "large TA" group (pTiTm4 and pTiHml); they correspond to the limited-and wide-host-range Ti plasmids, respectively (15, 41).The TB regions have an average size of about 20 kb and carry two auxin genes, TB-iaaM and TB-iaaH, together responsible for indoleacetic acid synthesis in the tumor cells (3,11,41)

Insertion Sequences

“…For two insertions of the Agrobacterium vitis element IS870, it was possible to determine the target sequence prior to insertion. This sequence was found to carry a single CTAG copy (103), an observation which remains consistent with a simple TA dinucleotide target duplication. For coherence, we have therefore assumed that all members of this family generate an identical (TA) target duplication and have therefore defined the tips of the elements accordingly (Fig.…”

“…The Tpase gene of several elements does not possess a termination codon. These include IS240C, a member of the IS6 family (57a); two members of the IS5 family, IS427 (77) and ISMk1 (228); and various members of the IS630 family, including IS870 and ISRf1 (103). Instead, some of these elements insert into a relatively specific target sequence in which the target DR produced on insertion itself generates the Tpase termination codon (see "IS630 family" below).…”

Temperature-responsive genetic loci in the plant pathogen Pseudomonas syringae pv. glycinea The GenBank accession numbers for the nucleotide sequences of mutants 560, 561, 562, 563, 564, 568, 570, 574, 590, 591, 593, 596, 599, 601, 605, 608, 613, 617, 618, 626 and 632 determined in this work are AF274322–AF274342, respectively.

“…3). The cloned DNA of mutant 562 showed strong sequence similarity to IS870, an insertion element frequently found in P. syringae and other plant pathogens (Fournier et al, 1993). Turner et al (1990) reported on the temperature sensitivity of transposition of class II transposons with optimal levels of transposition at room temperature or 30 mC versus 37-42 mC.…”

Analysis of Pseudomonas putida alkane-degradation gene clusters and flanking insertion sequences: evolution and regulation of the alk genes The EMBL accession numbers for the sequences reported in this paper are AJ245436 [P. putida (oleovorans) GPo1 alk gene clusters and flanking DNA], AJ233397 (P. putida P1 alk gene clusters and flanking DNA), AJ249793 (P. putida P1 nahKJ genes), AJ249825 [P. putida (oleovorans) GPo1 16S RNA gene] and AJ271219 (P. putida P1 16S RNA gene).

“…ISPpu1 is located immediately downstream of alkL in GPo1, and is related to the Agrobacterium tumefaciens and Agrobacterium vitis IS870 copies (Fournier et al, 1993). The IS element , the closest relative in the IS database is listed as well.…”