Functional Central Polypurine Tract Provides Downstream Protection of the Human Immunodeficiency Virus Type 1 Genome from Editing by APOBEC3G and APOBEC3B

Abstract

Lentiviruses utilize two polypurine tracts for initiation of plus-strand viral DNA synthesis. We have examined to what extent human immunodeficiency virus type 1 plus-strand initiation at the central polypurine tract (cPPT) could protect the viral genome from DNA editing by APOBEC3G and APOBEC3B. The presence of a functional cPPT, but not of a mutated cPPT, extensively reduced editing by both APOBEC3G and APOBEC3B of sequences downstream, but not upstream, of the cPPT, with significant protection observed as far as 400 bp downstream. Thus, in addition to other potential functions, the cPPT could help protect lentiviruses from editing by cytidine deaminases of the APOBEC family.Retroviral replication requires reverse transcription of particle-associated genomic viral RNA into double-stranded DNA. Reverse transcription follows sequential steps, during which discrete segments of DNA are synthesized by reverse transcriptase, with each of these segments being initiated at a precise site by a specific primer (30). Synthesis of the first (minus) strand of DNA uses genomic viral RNA as a template and is initiated by a tRNA primer that defines the 3Ј boundary of the U5 region of the long terminal repeat (LTR). Synthesis of the second (plus) strand of DNA uses minus-strand DNA as a template and is initiated by a short RNA primer corresponding to a purine-rich sequence, the polypurine tract (PPT) (24,27). While all retroviruses carry a PPT near the 3Ј end of the retroviral genome, defining the 5Ј boundary of the U3 region of the LTR, several retroviruses, including lentiviruses, carry a second PPT near the center of the genome. Central polypurine tracts (cPPT) have mainly been described for lentiviruses, including visna virus, feline immunodeficiency virus, bovine immunodeficiency virus, equine infectious anemia virus, simian immunodeficiency virus

Residual HIV-1 DNA Flap-independent nuclear import of cPPT/CTS double mutant viruses does not support spreading infection

“…Although this defect may vary in its amplitude depending on the nuclear import assay and the MOI used, it is consistently observed. Of note, central (+) strand initiation may also carry further benefits for HIV-1 replication besides assisting viral nuclear import, such as protection from APOBEC3G/F editing as previously shown [14,16]. Here, we show that even the most disrupted DNA Flap mutants still maintain residual nuclear import, but that this does not support spreading infection in human lymphocytic cells.…”

Mechanism of Enhanced HIV Restriction by Virion Coencapsidated Cytidine Deaminases APOBEC3F and APOBEC3G

“…Despite A3F-induced mutations being increased in the presence of A3G (Table 1), the reason for this did not appear to be due to an increase in the processive search mechanism of A3F. Another mechanism by which A3-induced mutations can be increased is if reverse transcription was slowed down, which would leave minus-strand DNA single stranded for a longer time (56)(57)(58). It is known that both A3G and A3F can decrease reverse transcriptase efficiency, which results in less late reverse transcript (LRT) formation and less proviral integration (36,37,(59)(60)(61)(62)(63)(64)(65)(66)(67)(68).…”

Viral and Cellular Requirements for the Nuclear Entry of Retroviral Preintegration Nucleoprotein Complexes

“…DNA plus-strand extension from the cPPT primer likely decreases the overall duration of reverse transcription within the cell [90,92], which may indirectly influence viral nuclear import during instances of limiting nucleotide concentrations, for example. Such a kinetic advantage to reverse transcription conferred by the cPPT is consistent with its ability to reduce the time-frame in which the viral single-stranded DNA is sensitive to the inhibitory activity of APOBEC3 cytosine deaminase restriction factors [93,94,95], and may similarly protect the RTC/PIC from other host defense proteins that could derail its trafficking [96]. …”