Abortive Gap Repair: Underlying Mechanism for Ds element Formation

Abstract

The mechanism by which the maize autonomous Ac transposable element gives rise to nonautonomous Ds elements is largely unknown. Sequence analysis of native maize Ds elements indicates a complex chimeric structure formed through deletions of Ac sequences with or without insertions of Ac-unrelated sequence blocks. These blocks are often flanked by short stretches of reshuffled and duplicated Ac sequences. To better understand the mechanism leading to Ds formation, we designed an assay for detecting alterations in Ac using transgenic tobacco plants carrying a single copy of Ac. We found frequent de novo alterations in Ac which were excision rather than sequence dependent, occurring within Ac but not within an almost identical Ds element and not within a stable transposase-producing gene. The de novo DNA rearrangements consisted of internal deletions with breakpoints usually occurring at short repeats and, in some cases, of duplication of Ac sequences or insertion of Ac-unrelated fragments. The ancient maize Ds elements and the young Ds elements in transgenic tobacco showed similar rearrangements, suggesting that Ac-Ds elements evolve rapidly, more so than stable genes, through deletions, duplications, and reshuffling of their own sequences and through capturing of unrelated sequences. The data presented here suggest that abortive Ac-induced gap repair, through the synthesis-dependent strand-annealing pathway, is the underlying mechanism for Ds element formation.Dissociation, or Ds, is the first discovered transposable element (TE). It was identified as a maize locus on chromosome 9, where breaks occur in the presence of Activator (Ac), a second gene found at a separate locus (33,34). Subsequent studies showed that Ac can transpose autonomously whereas Ds moves only in the presence of Ac (35,36). In addition, Ac activity can turn into a Ds type of instability, while no occurrences were found of Ds turning into Ac (37, 38). On the basis of these observations, McClintock proposed that Ds nonautonomous elements are derived from Ac through mutations (39). The proposal that Ac and Ds are phylogenetically related has been supported by molecular analysis, as described below, but the mechanism responsible for the conversion of Ac into Ds is still unknown.Ac is a 4.6-kb-long element flanked by 11-bp terminal inverted repeats (TIRs) (12). It encodes an 807-amino-acid protein, the transposase, necessary for both Ac and Ds transposition (28). Ds elements, on the other hand, do not encode a functional transposase but retain regions which are essential for their transposition (6). There are six fully sequenced Ds elements, all of which share with Ac nearly identical TIRs and fall into the following four categories: (i) those with nearly no similarity to Ac, like Ds1 (57); (ii) elements with highly similar subterminal regions but with internal deletions, like Ds9 (46); (iii) double Ds elements where one internally deleted Ds is inserted into another identical Ds in an inverted orientation (9); and (iv) Ds elements that contain bot...

State IIDissociationElement Formation FollowingActivatorExcision in Maize

“…Using these methods we identified 15 new Ds elements derived from Ac insertions at eight different sites in the maize genome. Sequence analysis revealed significant differences between these 15 Ds elements and previously reported Ds derivatives (Rubin and Levy 1997;Yan et al 1999). Specifically, several of these Ds elements lack direct repeats flanking the deletion junctions in the donor Ac.…”

“…For instance, Ds1-like elements contain large internal deletions sharing only the 59 terminal 13 bp and 39 terminal 26 bp of sequence with Ac (Sutton et al 1984). The Ds9 element is identical to Ac with the exception of a 194-bp simple deletion in the element (Rubin and Levy 1997). Other Ds family members, such as wxB4, share the TIR and subterminal sequence with Ac but carry internal sequences with no homology to Ac (Varagona and Wessler 1990).…”

“…Two related mechanisms have been proposed to account for the formation of Ds elements from Ac. Rubin and Levy (1997) concluded that state II Ds elements are generated through an abortive gap-repair process following Ac excision. However, in some instances, a synthesis-dependent strand-annealing (SDSA) repair mechanism is initiated at the time of this abortive gap repair.…”

“…However, in some instances, a synthesis-dependent strand-annealing (SDSA) repair mechanism is initiated at the time of this abortive gap repair. In these instances, polymerase slippage during DNA synthesis at the site of direct repeats within Ac may result in the insertion of sequences known as filler DNA at the deletion junction (Rubin and Levy 1997). Yan et al (1999) proposed that Ds elements were formed through slip mispairing during DNA synthesis resulting both in an internal deletion and in some cases the insertion filler DNA at the deletion junction (Yan et al 1999).…”

Diverse DNA transposons in rotifers of the class Bdelloidea

“…1 A Top). Deletion derivatives of DNA transposons are frequently observed and are thought to occur during abortive gap repair at the excision site, by means of aborted synthesis-dependent strand annealing and subsequent nonhomologous end-joining via microhomologies within the element (25,26).…”