General Information/What Is an IS?

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Classical IS

The original definition of an IS (Fig.3.1) was: a short, generally phenotypically cryptic, DNA segment encoding only the enzymes necessary for its transposition and capable of repeated insertion into many different sites within a genome using mechanisms independent of large regions of DNA homology between the IS and target [1][2][3]. Classical IS are between 0.7 and 2.5 kb in length, genetically compact with one or two open reading frames (orfs) which occupy the entire length of the IS and terminate in flanking imperfect terminal repeat sequences (IR) (Table 1). The orfs include the Tpase that catalyzes the DNA cleavages and strand transfers leading to IS movement and, in some cases, regulatory proteins. Their highly compact nature is illustrated by the fact that some IS have developed “recoding” strategies such as Programmed Ribosomal Frameshifting (involving ribosome slippage) and Programmed Transcriptional Realignment (involving RNA polymerase slippage) [4][5][6][7][8][9][10][11].

These permit assembly of different functional protein domains, effectively encoding two proteins of different functions in one DNA segment. IS also often generates a short flanking directly repeated duplication (DR) of the target DNA on insertion. These characteristics are not limited to prokaryotic IS but are also shared with most eukaryotic DNA transposons. Classical IS generally transpose using a double-strand DNA intermediate. However, for prokaryotic IS, this strict definition has been broadened over the years with the discovery of an increasing number of non-canonical derivatives and variants, some of which are described in the following sections. Moreover, as we learn more about diversity from sequenced genomes, classification is becoming more problematic because the large degree of MGE diversity is obscuring the borders between certain types of TE (see "Fuzzy Borders") [5]. Despite their abundance and diversity, the number of different chemical mechanisms used in TE movement is surprisingly limited, and many quite divergent TE share a similar mechanism.

Fig.3.1. What is an Insertion Sequence? Terminal inverted repeats (IRL and IRR) are shown as two-colored boxes (a and b) with functions for transposase binding (a) and recognition for cleavage and strand transfer (a). A single (left) or double (right) open reading frame is shown underneath the IS (blue arrow). The transposase of the IS on the right is produced by programmed -1 translational frameshifting. The reading frames are indicated within the IS. The product of the upstream frame generally acts as a regulatory protein. The indigenous Tpase promoter is shown located (by convention) in IRL. XXX and YYYY represent the short direct target repeat sequence which is generally duplicated during the insertion event.

Characteristics of insertion sequence families

Table 1. Abbreviations: DR, duplication repeat; IS, insertion sequence; ORF, open reading frame.
Families Sub-Groups Typical size-range (bp) DR (bp) Ends IRs No ORFs Frameshift Catalytic residues Mechanism
IS1 740–1180 8–9 GGnnnTG Y 2 ORFAB DDE copy-and-paste and cointegrate
single ORF 800–1200 0–9 N 1
ISMhu11 900–4600 0–10 Y 2 ORFAB
IS1595 ISPna2 1000–1150 8 GGCnnTG Y 1 DDNK copy-and-paste (?)
ISPna2+pass 1500–2600 8 1+pass
IS1016 700–745 7–9 GGGgctg DDEK
IS1595 900–1100 8 CcTGATT DDNK+ER4R7
ISSod11 1000–1100 8 nnnGcnTATC DDHK+ER4R7
ISNwi1 1080–1200 8 ggnnatTAT DDEK+ER4
ISNwi1+pass 1750–4750 8 1+pass
ISNha5 3450–7900 8 CGGnnTT 1 DDER/K
IS3 IS150 1200–1600 3–4 TG Y 2 ORFAB DDE copy-and-paste
IS407 1100–1400 4 TG
IS51 1000–1400 3–4 TG
IS3 1150–1750 3–4 TGa/g
IS2 1300–1400 5 TG
IS481 950–1300 4–15 TGT Y 1 DDE copy-and-paste (?)
IS1202 1400–1700 5 TGT Y 1 DDE
IS4 IS10 1200–1350 9 CT Y 1 DDE hairpin intermediate cut-and-paste
IS50 1350–1550 8–9 C hairpin intermediate
ISPepr1 1500–1600 7–8 -T-AA ?
IS4 1400–1600 10–13 -AAT ?
IS4Sa 1150–1750 8–10 CA ?
ISH8 1400–1800 10 ?
IS231 1450–5400 10–12 CAT 1 or + * *passenger genes
IS701 1400–1550 4 Y 1 DDE
ISH3 1225–1500 4–5 C-GT Y 1 DDE
IS1634 1500–2000 5–6 C Y 1 DDE
IS5 IS903 950–1150 9 GG Y 1 DDE
ISL2 850–1200 2–3
ISH1 900–1150 8 -GC
IS5 1000–1500 4 Ga/g
IS1031 850–1050 3 GAa/g
IS427 800–1000 2–4 Ga/g 2 ORFAB
IS1182 1330–1950 0–60 Y 1 DDE
IS6 700–900 8 GG Y 1 DDE co-integrate
IS21 1750–2600 4–8 TG Y 2 * DDE
IS30 1000–1700 2–3 Y 1 DDE copy-and-paste
IS66 2000–3000 8–9 GTAA Y 3* DDE*
ISBst12 1350–1900 1 DDE
IS256 1200–1500 8–9 Ga/g Y 1 DDE copy-and-paste
IS1249 1300 0–10 GG
ISC1250 1250 0–9 GG
ISH6 1450 8 GGT Y 1 DDE
ISLre2 1500–2000 9 Y 1 DDE
ISKra4 ISAzba1 1400–2900 0 Y 1 or + * DDE
ISMich2 1250–1400 8 GGG 1 or 2 ORFAB
ISKra4 1400–3700 9 GGG 1 or + *
IS630 1000–1400 2* Y 1 or 2 ORFAB DDE cut-and-paste
IS982 1000 3–9 AC Y 1 DDE
IS1380 1550–2000 4–5 CC Y 1 DDE
ISAs1 1200–1500 8–10 CAGGG Y 1
ISL3 1300–2300 8 GG Y 1
Tn3 >3000 0 GGGG Y >1 DDE co-integrate
ISAzo13 1250–2200 0–4 Ga/g Y 1
IS110 1200–1550 0 N 1 DEDD
IS1111 Y*
IS91 1500–2000 0 N 1 HUH/Y2 rolling-circle
IS200/IS605 IS200 600–750 0 0 1* HUH/Y1 peel-and-paste
IS605 1300–2000 2* HUH/Y1**
IS607 1700–2500 0 N 2* Serine**
ISNCY * IS892 1600 0–8 CTAG Y 2 ORFAB
ISLbi1 1400–1500 5 Y 1
ISMae2 1400–2400 9 CAG Y 1
ISPlu15 800–1000 0 N 1
ISA1214 1000–1200 8–12 Y 2
ISC1217 1200 6–8 TAG Y 1
ISM1 1300–1600 8–9 Y 1
IS1202 1400–1700 5 TGT Y 1 DDEQ
ISDol1 1600–1900 6–7 Y 1 DDE
* ISNCY = Insertion Sequence Not Classified Yet

New types of IS

One example of this expanding diversity is the identification of another entire class of IS [12][13][14]. Members of this class use an entirely different mechanism of transposition involving single-strand circular DNA intermediates which appear to target stalled replication forks [15] (Fig.3.2). They possess small transposases (~150 aa) which are completely different to the classical IS in the type of chemistry they catalyze (Groups with HUH Enzymes). Another example are the casposons which are related to CRISPRs but whose transposition has yet to be fully characterized [16][17][18][19].

Fig.3.2. Organization of the IS200/IS605 family insertion sequences that transpose using single-strand DNA intermediates. The top of the figure shows the genetic organization. The generic IS is shown as a box. The element-specific tetra- (IS608) or penta- (ISDra2) nucleotide target site is shown boxed in red on the left. Left (LE) and right (RE) ends carrying subterminal hairpins are presented as red boxes. The transposase reading frame, tnpA, is shown within the IS as a red line and the accessory frame tnpB as a blue line. The direction of transcription is indicated by the arrowhead. The bottom of the figure shows the subterminal secondary structures at the left (red) end LE and the right (blue) end RE. The left and right cleavage sites are presented as black The blue arrows indicate the interaction between the cleavage sites and the "guide" sequences at the foot of the secondary structures necessary for the cleavage reactions.


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