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Multivalent interactions essential for lentiviral integrase function

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journal contribution
posted on 06.05.2022, 08:16 authored by Allison Ballandras-Colas, Vidya Chivukula, Dominika T Gruszka, Zelin Shan, Parmit K Singh, Valerie E Pye, Rebecca K McLean, Gregory J Bedwell, Wen Li, Andrea Nans, Nicola J Cook, Hind J Fadel, Eric M Poeschla, David J Griffiths, Javier Vargas, Ian A Taylor, Dmitry Lyumkis, Hasan Yardimci, Alan N Engelman, Peter Cherepanov
A multimer of retroviral integrase (IN) synapses viral DNA ends within a stable intasome nucleoprotein complex for integration into a host cell genome. Reconstitution of the intasome from the maedi-visna virus (MVV), an ovine lentivirus, revealed a large assembly containing sixteen IN subunits1. Herein, we report cryo-EM structures of the lentiviral intasome prior to engagement of target DNA and following strand transfer, refined at 3.4 and 3.5 Å resolution, respectively. The structures elucidate details of the protein-protein and protein-DNA interfaces involved in lentiviral intasome formation. We show that the homomeric interfaces involved in IN hexadecamer formation and the α-helical configuration of the linker connecting the C-terminal and catalytic core domains are critical for MVV IN strand transfer activity in vitro and for virus infectivity. Single-molecule microscopy in conjunction with photobleaching reveals that the MVV intasome can bind a variable number, up to sixteen molecules, of the lentivirus-specific host factor LEDGF/p75. Concordantly, ablation of endogenous LEDGF/p75 results in gross redistribution of MVV integration sites in human and ovine cells. Our data confirm the importance of the expanded architecture observed in cryo-EM studies of lentiviral intasomes and suggest that this organization underlies multivalent interactions with chromatin for integration targeting to active genes.

Funding

Crick (Grant ID: 10061, Grant title: Cherepanov FC001061) Crick (Grant ID: 10015, Grant title: STP Structural Biology) Crick (Grant ID: 10178, Grant title: Taylor,I FC001178) Crick (Grant ID: 10221, Grant title: Yardimci FC001221)

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