Structural basis of seamless excision and specific targeting by piggyBac transposase

• Published in: Nature communications Vol. 11; no. 1; p. 3446
• Main Authors: Chen, Qiujia, Luo, Wentian, Veach, Ruth Ann, Hickman, Alison B., Wilson, Matthew H., Dyda, Fred
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group 10.07.2020; Nature Publishing Group UK; Nature Portfolio
• Subjects: Asymmetry; Binding sites; Complex formation; Deoxyribonucleic acid; Dimers; DNA; DNA structure; Domains; Footprints; Genome editing; Genomes; Intermediates; Molecular structure; Transposase; Transposition; Transposons
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-020-17128-1

Abstract The piggyBac DNA transposon is used widely in genome engineering applications. Unlike other transposons, its excision site can be precisely repaired without leaving footprints and it integrates specifically at TTAA tetranucleotides. We present cryo-EM structures of piggyBac transpososomes: a synaptic complex with hairpin DNA intermediates and a strand transfer complex capturing the integration step. The results show that the excised TTAA hairpin intermediate and the TTAA target adopt essentially identical conformations, providing a mechanistic link connecting the two unique properties of piggyBac . The transposase forms an asymmetric dimer in which the two central domains synapse the ends while two C-terminal domains form a separate dimer that contacts only one transposon end. In the strand transfer structure, target DNA is severely bent and the TTAA target is unpaired. In-cell data suggest that asymmetry promotes synaptic complex formation, and modifying ends with additional transposase binding sites stimulates activity.