B-to-A transition in target DNA during retroviral integration

Abstract

Integration into host target DNA (tDNA), a hallmark of retroviral replication, is mediated by the intasome, a multimer of integrase (IN) assembled on viral DNA (vDNA) ends. To ascertain aspects of tDNA recognition during integration, we have solved the 3.5 Å resolution cryo-EM structure of the mouse mammary tumor virus (MMTV) strand transfer complex (STC) intasome. The tDNA adopts an A-like conformation in the region encompassing the sites of vDNA joining, which exposes the sugar-phosphate backbone for IN-mediated strand transfer. Examination of existing retroviral STC structures revealed conservation of A-form tDNA in the analogous regions of these complexes. Furthermore, analyses of sequence preferences in genomic integration sites selectively targeted by six different retroviruses highlighted consistent propensity for A-philic sequences at the sites of vDNA joining. Our structure additionally revealed several novel MMTV IN-DNA interactions, as well as contacts seen in prior STC structures, including conserved Pro125 and Tyr149 residues interacting with tDNA. In infected cells, Pro125 substitutions impacted the global pattern of MMTV integration without significantly altering local base sequence preferences at vDNA insertion sites. Collectively, these data advance our understanding of retroviral intasome structure and function, as well as factors that influence patterns of vDNA integration in genomic DNA.

Mechanisms of HIV-1 Integrase Resistance to Dolutegravir and Potent Inhibition of Drug Resistant Variants

“…Each step of DNA integration, up to formation of the integration intermediate, occurs within a series of stable nucleoprotein complexes (intasomes), involving a multimer of IN and a pair of viral DNA ends. The atomic structures have now been determined for a variety of retroviral intasomes (4)(5)(6)(7)(8)(9)(10)(11)(12)(13), including HIV-1 (14,15), revealing differences in the oligomeric assemblies and how they engage target DNA. The formation of intasome species, and the intasome-mediated insertion of the vDNA into the host genome, are essential steps in the viral replication cycle [see (1,2) among the best therapeutic options for use in combination therapies to treat HIV-infected patients (18).…”

Unraveling the Palindromic and Non-Palindromic Motifs of Retroviral Integration Site Sequences by Statistical Mixture Models

“…Since there are no clear correlates of IN-tDNA interface composition and the strength of tDNA motifs, the motif may reflect a need for tDNA to adopt specific conformation. Yet, there is no major distortion of B-form tDNA structure at the position occupied by preferred T 18 . Also, the number of documented H-bond-forming interactions between IN and tDNA does not correlate with the preference for T-palindrome upstream to the STR.…”

“…One explanation for the nucleotide preferences might reside in specific interactions at the IN-tDNA interface. About 3 bp upstream to the STR site, structurally conserved intrusion into the minor groove of small amino acids like serine (HIV, RSV), proline (HTLV, MVV, MLV 36 , MMTV 18 ), and alanine (PFV) was documented. Albeit no nucleotide-specific interactions seem to be present at the position, mutations of HIV-1 IN S119 34,35,37 affect the preference of IN target site selection.…”

“…Globally, tethering to chromatin by cellular proteins drives the preferences [8][9][10][11][12][13] . Locally, the occupancy of nucleosomes and the physical attributes of DNA affect the target site selection [14][15][16][17][18] .…”

“…Typically, the STR sites on both strands of tDNA are 4 to 6 nucleotides apart depending on the retrovirus. Structural studies demonstrated that IN assembles into symmetric multimeric complexes that bend the tDNA enabling the STR inside a widened major groove of tDNA [18][19][20][21][22][23][24][25][26][27] .…”