Bouchonnet F, Lecossier D, Clavel F, Hance AJ; Conference on Retroviruses and Opportunistic Infections (11th : 2004 : San Francisco, Calif.).
Program Abstr Conf Retrovir Oppor Infect 11th 2004 San Franc Calif. 2004 Feb 8-11; 11: abstract no. 355.
INSERM U552, Paris, France
BACKGROUND: Incorporation of the cytosine deaminase APOBEC3G into HIV particles, which is inhibited by the Vif accessory protein, induces G to A mutations into viral plus-strand DNA. These mutations are thought to result from the introduction of C to U changes in newly synthesized minus-strand DNA, but alternatives have been proposed. In addition, APOBEC3G activity could participate in viral evolution, and this would be accelerated by error-prone repair of uracil residues in integrated DNA, as described in other cell types. To rule out editing mechanisms such as deamination of dCTP before incorporation into DNA or editing of DNA/RNA hybrids, and to search for error-prone repair of U residues, we have examined the pattern of hypermutation in HIV proviruses resulting from infection of H9 cells by Vif-defective HIV-1.METHODS: To specifically evaluate stably integrated viral sequences, high molecular weight DNA was extracted from clonal populations of H9 cells chronically infected with DeltaVif HIV-1. A 1193bp fragment of pol, which includes the central polypurine tract (PPT), was amplified, cloned and multiple clones were sequenced.RESULTS: Each cloned cell line contained multiple integrated proviruses. The 15 sequences contained 320 G to A substitutions, but only 28 other base substitutions. Importantly, only 4 of these other substitutions involved a G residue, and 3 of these G residues had not been subject to G to A substitution in any other sequence. Despite the favorable sequence context (multiple adjacent G residues including 5 consecutive "GGR motifs" identified as hot-spots for editing by APOBEC3G), the PPT was not edited in any sequence. In contrast, outside the PPT, 10/10 sequences containing 3 or more consecutive Gs had been edited, and 45/47 "GGR" triplets contained G to A transitions in one or more sequences (p<0.0001 by Fisher's exact test).CONCLUSIONS: The absence of editing of the polypurine tract supports the idea that APOBEC3G targets single-stranded minus-strand DNA, not dCTP or RNA/DNA hybrids. We found no evidence that error-prone repair of uracil residues is a source for further diversification.
Publication Types:
Keywords:
- Base Sequence
- Communicable Diseases
- DNA, Viral
- Gene Products, vif
- Genes, pol
- HIV
- HIV Infections
- HIV-1
- Proviruses
- RNA
- Reverse Transcription
- genetics
- reverse transcriptase, Human immunodeficiency virus 1
Other ID:
UI: 102271315
From Meeting Abstracts