An in vivo transposase-catalyzed single-stranded DNA circularization reaction.
Open Access
- 15 November 1995
- journal article
- Published by Cold Spring Harbor Laboratory in Genes & Development
- Vol. 9 (22) , 2846-2858
- https://doi.org/10.1101/gad.9.22.2846
Abstract
Expression of the bacterial insertion sequence IS911 transposase in vivo leads to excision and circularization of IS911-based transposons. We show here that transposase produces an unusual molecular form generated by single-strand cleavage, transfer, and ligation of one end of the element to the opposite end. When the transposon is carried by a circular plasmid, this results in the formation of a "figure-eight" molecule in which a single strand of the transposon is circularized while the corresponding strand of the vector backbone retains a single-strand interruption at this position. The results show that a 3' end of the transposon is transferred to the opposite target end. Transposase is therefore capable of introducing single-strand cleavages at the ends of the element, an activity similar to that of retroviral integrases with which it shares significant similarities in amino acid sequence. Kinetic studies demonstrate that the figure-eight accumulates earlier than transposon circles after transposase induction and disappears before circles after inhibition of transposase expression, raising the possibility that the figure-eight molecules are precursors to the circles. Therefore, IS911 excision as a circle may not occur by double-strand cleavage leading to its prior separation from the vector backbone in a linear form but could proceed by consecutive circularization of each strand.Keywords
This publication has 33 references indexed in Scilit:
- Translational Control in Production of Transposase and in Transposition of Insertion Sequence IS3Journal of Molecular Biology, 1994
- Retroviral DNA integration: lessons for transposon shufflingGene, 1993
- Tn7 transposition: Target DNA recognition is mediated by multiple Tn7-encoded proteins in a purified in vitro systemPublished by Elsevier ,1993
- Elimination of Tec elements involves a novel excision process.Genes & Development, 1993
- Circular forms of developmentally excised DNA in Euplotes crassus have a heteroduplex junction.Genes & Development, 1993
- Programmed translational frameshifting and initiation at an AUU codon in gene expression of bacterial insertion sequence IS911Journal of Molecular Biology, 1991
- Kinetic and structural analysis of a cleaved donor intermediate and a strand transfer intermediate in Tn10 transpositionCell, 1991
- Developmentally coordinated en masse excision of a highly repetitive element in E. crassusCell, 1989
- Detection of circular forms of eliminated DNA during macronuclear development in E. crassusCell, 1989
- An essential replication gene, repA, of plasmid pSC101 is autoregulatedJournal of Molecular Biology, 1985