Insertion sequence (IS) elements are small (0.75 to 1.5 kbp) transposable elements found in bacteria (1). They usually encode only a transposase gene and special sequences at the tips of the element on which the recombinase acts to move the element from place to place. IS elements thus structurally resemble, for example, P elements from Drosophila, the widespread Tc1/mariner elements, and Ac elements from maize, although IS elements are generally much smaller. Like P, Tc1/mariner, and Ac elements, IS elements alter DNA in the host genome upon insertion and can cause gene inactivation.
IS elements were among the first nonmaize transposable elements discovered, being identified in bacteria in the late 1960s as the cause of highly polar mutations in bacterial operons and also as components of translocatable antibiotic-resistance segments. It was possible to manipulate bacterial
DNA molecules at this time because of their relatively small size, something that was not possible for other organisms such as maize, and the direct characterization of the DNA of these polar mutations revealed that they resulted from an insertion of a discrete piece of DNA.
Some IS elements also associate to form what are called composite elements in which two ISs flank a region of DNA that encodes a selectable trait, for example an antibiotic-resistance determinant (Fig. 1). The action of transposase at the two external edges of the IS elements can move the entire IS-drug resistance gene-IS element from place to place (Fig. 1). Each IS element can also move independently, although this does not result in movement of the antibiotic resistance determinant. IS elements are found as individuals at various positions in bacterial chromosome.
Figure 1. IS elements can associate to form composite transposons. IS elements encode information to promote their translocation, a transposase (arrow) and special sequences at their ends (triangles). Composite transposons are formed by two IS elements flanking a drug resistance determinant, here tetracycline. When the inside (I) and outside (O) ends of an individual IS element are the substrates for recombination, the IS element alone translocates. When the outside (O) ends of the two ISs flanking the antibiotic resistance determinant are the substrates for recombination, the entire IS-drug resistance-IS segment translocates.
Particularly well-studied IS sequences are IS10 (2) and IS50 (3). The composite transposon Tn10 consists of two IS10 elements flanking a DNA segment that encodes a tetracycline-resistance determinant. The composite transposon Tn5 is composed of two IS50 elements flanking a kanamycin-resistance encoding segment. Both of these elements translocate by a cut-and-paste mechanism, in which the element is excised from the donor DNA and then inserts into the target DNA. They and their derivatives have also been invaluable reagents in the genetic analysis of bacteria (4). The strategy of using transposon mutagenesis to tag genes physically to allow gene isolation has been a very valuable technique.
Another class of IS elements—members of the IS3 family that includes IS3 and IS911—have an interesting strategy for expressing transposase. The element contains two open reading frames, OrfA and OrfB. OrfA encodes the DNA-binding determinant that directs this protein to the ends of the transposon, but OrfA has no transposase activity. The actual transposase OrfAB results from a frameshifting event near the end of OrfA that joins OrfA and OrfB (5). The amount of transposase, and thus the frequency of transposition, is determined by the amount of frameshifting. These elements also appear to transpose by an unusual mechanism (6, 7). Transposition begins by cleavage at one end of the transposon, exposing a 3′OH end of the element, followed by an intramolecular attack of that 3′OH end to just outside the 5′ end of that same strand; note that this is the same chemistry as is involved in other transposition reactions, the only difference being that the joining reaction is intramolecular, rather than intermolecular. By either another such single-strand cleavage and intramolecular joining, or by replication, a double-stranded DNA circle results in which the IS ends are closely juxtaposed, with just a few nucleotides separating them. This "circle junction" version of the IS element then interacts with the target site and is inserted by breakage at 3′ ends of the transposon and joining the exposed ends to the target DNA.
It should be appreciated that although some of the well-studied elements and widely used bacterial elements such as Tn10 and Tn5 contain IS elements, certainly not all bacterial transposable elements contains IS sequences. Tn3, Mu phage, and Tn7 are all examples of bacterial elements that lack IS sequences, and these elements do not associate to form composite transposons.
