Repair of oxidative DNA base damage in the host genome influences the HIV integration site sequence preference.

Geoffrey R Bennett,Kristine E Yoder,Robert W Sobol,Ryan Peters,Xiao-Hong Wang,Richard Fishel,Jeungphill Hanne,Ralf Bundschuh,Maria Spies

doi:10.1371/journal.pone.0103164

Abstract

Host base excision repair (BER) proteins that repair oxidative damage enhance HIV infection. These proteins include the oxidative DNA damage glycosylases 8-oxo-guanine DNA glycosylase (OGG1) and mutY homolog (MYH) as well as DNA polymerase beta (Polβ). While deletion of oxidative BER genes leads to decreased HIV infection and integration efficiency, the mechanism remains unknown. One hypothesis is that BER proteins repair the DNA gapped integration intermediate. An alternative hypothesis considers that the most common oxidative DNA base damages occur on guanines. The subtle consensus sequence preference at HIV integration sites includes multiple G:C base pairs surrounding the points of joining. These observations suggest a role for oxidative BER during integration targeting at the nucleotide level. We examined the hypothesis that BER repairs a gapped integration intermediate by measuring HIV infection efficiency in Polβ null cell lines complemented with active site point mutants of Polβ. A DNA synthesis defective mutant, but not a 5′dRP lyase mutant, rescued HIV infection efficiency to wild type levels; this suggeted Polβ DNA synthesis activity is not necessary while 5′dRP lyase activity is required for efficient HIV infection. An alternate hypothesis that BER events in the host genome influence HIV integration site selection was examined by sequencing integration sites in OGG1 and MYH null cells. In the absence of these 8-oxo-guanine specific glycosylases the chromatin elements of HIV integration site selection remain the same as in wild type cells. However, the HIV integration site sequence preference at G:C base pairs is altered at several positions in OGG1 and MYH null cells. Inefficient HIV infection in the absence of oxidative BER proteins does not appear related to repair of the gapped integration intermediate; instead oxidative damage repair may participate in HIV integration site preference at the sequence level.

Highlights

Retroviruses are defined by two enzymatic activities, reverse transcription and integration [1]
We have further explored the effects of base excision repair (BER) proteins on HIV integration with well-defined BER mutant cell lines
polymerase beta (Polb) null murine embryonic fibroblasts were complemented with an empty vector or the wild type Polb cDNA (Figure 2A, B) [33]

Summary

Introduction

Retroviruses are defined by two enzymatic activities, reverse transcription and integration [1]. The viral enzyme reverse transcriptase copies the viral genomic RNA to a linear double stranded cDNA. The cDNA is part of a pre-integration complex (PIC) that is poorly understood. The PIC travels to the nucleus where the viral protein integrase mediates the covalent joining of the viral cDNA to the host chromosome. The two ends of the viral cDNA are joined to the host DNA 4–6 base pairs apart, depending on the retrovirus. The process of integration yields an integration intermediate comprised of the viral cDNA flanked by 5 base gaps of host sequence and 59 dinucleotide flaps of viral sequence. Host enzymes are assumed to repair this integration intermediate, but identities of the specific repair proteins remain unknown [2]

Methods

Results

Conclusion