The cytidine deaminase CEM15 induces hypermutation in newly synthesized HIV-1 DNA

Abstract

High mutation frequency during reverse transcription has a principal role in the genetic variation of primate lentiviral populations. It is the main driving force for the generation of drug resistance and the escape from immune surveillance. G to A hypermutation is one of the characteristics of primate lentiviruses, as well as other retroviruses, during replication in vivo and in cell culture 1-6 . The molecular mechanisms of this process, however, remain to be clarified. Here, we demonstrate that CEM15 (also known as apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like 3G; APOBEC3G) 7,8 , an endogenous inhibitor of human immunodeficiency virus type 1 (HIV-1) replication, is a cytidine deaminase and is able to induce G to A hypermutation in newly synthesized viral DNA. This effect can be counteracted by the HIV-1 virion infectivity factor (Vif). It seems that this viral DNA mutator is a viral defence mechanism in host cells that may induce either lethal hypermutation or instability of the incoming nascent viral reverse transcripts, which could account for the Vif-defective phenotype. Importantly, the accumulation of CEM15-mediated non-lethal hypermutation in the replicating viral genome could potently contribute to the genetic variation of primate lentiviral populations.HIV-1 Vif protein is required for viral replication in vivo and in some 'non-permissive' cells, such as peripheral blood mononuclear cells, macrophages and H9 T cells 9-11 . The vifdefective viruses (Δvif) from non-permissive cells cannot complete reverse transcription, or the newly synthesized DNA cannot exist in the target cells for a significant time period 12-14 . Recently, it has been demonstrated that CEM15 is an endogenous inhibitor of HIV-1 that exists only in non-permissive cells. Its inhibitory effect on HIV-1 replication can be counteracted by HIV-1 Vif protein 7 . As Vif binds to HIV-1 RNA in the cytoplasm of virusproducing cells 15-17 , we investigated whether CEM15, which shares significant homology with some other cytidine deaminases that edit RNA, could also edit HIV-1 genomic or spliced RNA. We have sequenced the nearly full-length genomic RNA (>98%) of HIV-1 from the Δvif virions generated from H9 T cells by polymerase chain reaction with reverse transcription (RT-PCR) techniques (primer pairs are listed in Supplementary Table S1). Compared with the sequence of pNL4-3Δvif DNA, the change of genomic RNA in the virions is not significant. We have found an A to G change at positions 2257 and 3608, and a G to T change at position 9418 (data not shown). We have also sequenced several spliced HIV-1(NL4-3Δvif) RNA in H9 cells ( Supplementary Fig. S1 and Table S1); however, no mutations were demonstrated.Correspondence and requests for materials should be addressed to H.Z (hui.zhang@jefferson.edu).. Supplementary Information accompanies the paper on www.nature.com/nature. Competing interests statementThe authors declare that they have no competing financial interests. Table S1) that we have analysed (Fig. 1a, row 2 ...

Large-scale DNA editing of retrotransposons accelerates mammalian genome evolution

“…To detect edited elements, we assumed that a newly edited element should be almost identical to its ancestral element except for a few dense clusters of G-to-A mismatches, as such clusters are the hallmark of APOBEC3 editing [10][11][12][13] . To confirm that the mismatches are due to DNA editing, we exploited the asymmetry, or strand specificity, of editing: as opposed to random mutagenesis, APOBEC3 generates C-to-U mutations specifically on the negative strand of the element (with respect to the ORF), yielding clusters of G-to-A mutations, but not C-to-T, on the positive strand [10][11][12] .…”

“…To confirm that the mismatches are due to DNA editing, we exploited the asymmetry, or strand specificity, of editing: as opposed to random mutagenesis, APOBEC3 generates C-to-U mutations specifically on the negative strand of the element (with respect to the ORF), yielding clusters of G-to-A mutations, but not C-to-T, on the positive strand [10][11][12] . We therefore aligned, pairwise, the positive strands of all retrotransposons from selected families, and scanned the alignments for windows containing a particularly large number of G-to-A mismatches but no mismatches of any other type (Fig.…”

“…Occasionally, multiple cytosine-to-uracil deaminations are introduced on the negative-strand retroviral DNA after reverse transcription by proteins of the APOBEC3 (apolipoprotein B mRNA-editing enzyme and catalytic polypeptide 3) family [10][11][12][13] . While encounter with APOBEC3 impairs some viruses, others successfully integrate into the host genome, bearing G-to-A mutations compared with their original sequence.…”

A putative antiviral role of plant cytidine deaminases

“…Lethal mutagenesis through deamination of RNA/DNA by cytidine deaminases has been proven to work as an antiviral mechanism against retroviruses 16– 19, 23– 27 , and some DNA 28– 31 and RNA 32 viruses infecting mammals. Our results show that the A. thaliana CDA1 gene has some degree of mutagenic activity on the pararetrovirus CaMV genome.…”

Roles of HIV-1 auxiliary proteins in viral pathogenesis and host-pathogen interactions

“…The molecular nature of permissivity and the exact function of Vif in infection of nonpermissive cells was not known until recently when a series of reports showed that a host cellular protein known as APOBEC3G (apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3G) is a potent inhibitor of HIV infection in the nonpermissive cells [70,71]. APOBEC3G is a member of the cytidine deaminase family, which prevents viral cDNA synthesis via deaminating deoxycytidines (dC) in the minus-strand retroviral cDNA replication intermediate [72][73][74][75][76]. As a result, it creates stop codons or G-A transitions in the newly synthesized viral cDNA which is then subjective to elimination by host DNA repair machinery [74,76].…”