# Retroviral intasome architecture shapes the dynamics of target DNA search and integration

## TL;DR

This study compares the behavior of intasomes from two retroviruses, showing how their structure affects DNA integration speed and efficiency.

## Contribution

The first single-molecule analysis of an orthoretrovirus intasome's interaction with target DNA.

## Key findings

- MMTV intasomes remain active for ~80 minutes, much longer than PFV intasomes which plateau in ~2 minutes.
- MMTV intasomes integrate more efficiently with manganese than magnesium and search DNA more slowly than PFV intasomes.
- The time between integrating the two viral DNA ends is ~3 times slower in MMTV compared to PFV intasomes.

## Abstract

Recombinant retroviral intasomes assembled from purified integrase (IN) and oligonucleotides mimicking viral DNA ends (vDNA) faithfully recapitulate concerted integration in vitro. Structural studies of retroviral intasomes have revealed an array of IN oligomer forms, which appear to share a conserved intasome core coordinating the vDNA ends for strand transfer into target DNA. Here we have explored the biochemical and dynamic properties of the mouse mammary tumor virus (MMTV) octameric intasome. We show that MMTV intasomes continue to accumulate concerted integration products for ~80 min in vitro, whereas prototype foamy virus (PFV) intasomes plateau within ~2 min. MMTV integration activity peaks within the range of physiological ionic strength and is more active in the presence of manganese compared to magnesium. Single-molecule images demonstrate that the target DNA search by MMTV intasomes appears rate-limiting, similar to PFV intasomes. The time between strand transfer of the two MMTV vDNA ends into the target DNA is ~ 3 fold slower than PFV intasomes. This is the first report of the dynamics of an orthoretrovirus intasome interacting with target DNA with single molecule resolution.

Retroviruses insert their genetic material into the DNA of infected cells, a process carried out by a viral enzyme called integrase. Integrase molecules assemble with viral DNA ends to form a complex known as the intasome, which joins the viral DNA to the host DNA. Although the overall steps of integration are shared among retroviruses, the details of how different intasomes find and react with host DNA have remained unclear. Here, we characterized the intasome from the mouse mammary tumor virus (MMTV) for comparison to the prototype foamy virus (PFV) intasome. Using biochemical and single-molecule imaging approaches, we found that the larger MMTV intasome remains catalytically active longer than the smaller PFV complex and moves more slowly while searching for a target site on DNA. We found a common mechanism for how the intasomes interact with target DNA, while the details appear to vary based on the size of the complex.