DEFINITION
APOBEC3G (apolipoprotein B mRNA-editing, enzyme-catalytic, polypeptide-like 3G) is a human enzyme encoded by the APOBEC3G gene that belongs to the APOBEC superfamily of proteins. This family of proteins has been suggested to play an important role in innate anti-viral immunity. APOBEC3G is a cytadine deaminase enzyme and is therefore known as DNA dC->dU-editing enzyme APOBEC-3G and APOBEC-related cytidine deaminase (ARCD).
THE GENE
CHEMICAL STRUCTURE AND IMAGES
APOBEC3G has a symmetric structure, resulting in 2 homologous catalytic domains, the N-terminal (CD1) and C-terminal (CD2) domains, each of which contains a Zn2+ coordination site. Each domain also has the typical His/Cys-X-Glu-X23–28-Pro-Cys-X2-Cys motif for cytidine deaminases. However, unlike the typical cytidine deaminases, APOBEC3G contains a unique alpha helix between two beta sheets in the catalytic domain that could be a cofactor binding site. (Fig. 1)
Fig. 1
CD2 is catalytically active and vital for deamination and motif specificity. CD1 is catalytically inactive, but very important for binding to DNA and RNA and is key to defining the 5’->3’ processivity of APOBEC3G deamination. Interestingly, CD2 is unable to exert deaminase activity without the presence of CD1. Native [APOBEC3G] is composed of monomers, dimers, trimers, tetramers, and higher order oligomers. While it is thought that APOBEC3G functions as a dimer, it is possible that it actually functions as a mix of monomers and oligomers.
The D128 amino acid residue, which lies within CD1 (Fig. 1), appears to be particularly important for APOBEC3G interactions with Vif because a D128K point mutation prevents Vif-dependent depletion of APOBEC3G. Additionally, amino acids 128-130 on APOBEC3G form a negatively charged motif that is critical for interactions with Vif and the formation of APOBEC3G-Vif complexes. Furthermore, residues 124-127 are important for encapsidation of APOBEC3G into HIV-1 virions and the resulting antiretroviral activity.
Protein Aminoacids Percentage
CELLULAR FUNCTIONS
APOBEC deaminases-mutases with defensive roles for immunity. 2009
The Apo3G subfamily contains 7 members (Apo3A-H) and plays a significant role within the innate immune system. Many of the Apo3 proteins can restrict the replication of retroviruses and the mobility of retroelements. Apo3G is located into cytoplasm and was discovered in a subtractive hybridization screen as the cellular factor that blocks the replication of an HIV-1 strain that is deficient for its viral infectivity factor protein. The HIV-1 expresses its Vif protein to overcome the Apo3G imposed replication block primarily by binding to Apo3G and targeting it for polyubiquitylation and proteasomal degradation.
The antiretroviral enzyme APOBEC3G is degraded by the proteasome in response to HIV-1 Vif. 2003
Sequencing of reverse transcription products of Vif-deficient virions produced by permissive cells transfected with an hA3G-expressing vector revealed a hypermutation of HIV-1 DNA. Almost all mutations were G-to-A transitions when read on the plus DNA strand. Similar results were obtained when the products of endogenous or intracellular reverse transcription of Vif-deficient virions produced by nonpermissive cells were analyzed. These mutations are the result of cytosine deamination in the minus DNA strand, which convert cytosines into uracils, leading to G-to-A mutations in the plus strand. In contrast, hA3G does not edit HIV-1 genomic RNA. hA3G is catalytically active only on single-stranded DNA and not on DNA-DNA and DNA-RNA hybrids or on single-stranded RNA, protecting in this way genomic DNA (dsDNA) and mRNAs (ssRNA). Indeed, hA3G incorporated into virions is inhibited by HIV-1 genomic RNA and activated during plus-strand DNA synthesis as the RNA is degraded by the RNase activity of reverse transcriptase. The probability that a given minus-strand cytosine is deaminated depends on the time it remains single stranded and is increased by resistance mutations reducing the processivity of reverse transcriptase. hA3G proceeds processively from 3' to 5' of minus-strand DNA, and the dispersed hypermutation of the genome is the result of rapid transfers of hA3G between segments of the minus-strand HIV-1 DNA. Cytosine deamination by hA3G is not random. T G G G (as read in the plus strand) is the tetranucleotide that is most frequently modified in vitro and in cell culture, and the hot-spot consensus sequence of this enzyme is H G G R (where H is C, T, or A and R is A or G) (the mutated residue is highlighted).
Beyond HIV-1, some of the Apo3 enzymes inhibit other retroviruses such as simian immunodeficiency virus, murine leukemia virus and human T-cell leukemia virus type 1. Similar to retroviruses, HBV packages RNA into virions that must undergo a reverse transcription step to form the double-stranded DNA genome. This step allows Apo3G to disrupt HBV replication in hepatoma cell lines.
Inhibition of hepatitis B virus replication by APOBEC3G. 2004
Lastly, Apo3G was recently identified as the protein encoded by the Recovery from Friend virus 3 (Rfv3) gene, which influences retroviral neutralizing antibody responses and viremia in mice. In addition to the anti-viral properties, the APOBEC3 enzymes also protect the human genome against endogenous retrotransposons. The mobilization of these elements poses serious risks of cancers and genetic diseases. Therefore, APOBEC proteins provide a very important biological role by inhibiting the replication of these retroelements. For example Apo3G inhibits the yeast Ty1 LTR retrotransposon.
Inhibition of a yeast LTR retrotransposon by human APOBEC3 cytidine deaminases. 2005
APOBEC3G hypermutates genomic DNA and inhibits Ty1 retrotransposition in yeast. 2005
On the other hand, Apo3G does not inhibit LINE1 retrotranposition, but effectively inhibits LINE1-dependent retrotransposition of Alu retroelements without cytidine deamination activity.
High-molecular-mass APOBEC3G complexes restrict Alu retrotransposition. 2006
It is thought that Apo3G binding the Alu RNA keeps it sequestered in the cytoplasm away from the nuclear-located LINE1 machinery that is required for Alu retrotransposition.
SYNTHESIS AND TURNOVER
APOBEC3G mRNA is expressed in certain cells, referred to as non-permissive cells (cells where the viral cycle is not productive), in which HIV-1 cannot properly infect and replicate in the absence of Vif. Such cells include physiologically relevant primary CD4 T lymphocytes and macrophages.The encapsidation of APOBEC3G into HIV-1 virions is very important for the spread of APOBEC3G and the exertion of anti-retroviral activity.
Encapsidation of APOBEC3G may occur by at least the following four proposed mechanisms (Fig. 2):
1. Non-specific packaging of APOBEC3G
2. APOBEC3G interaction with host RNA
3. APOBEC3G interaction with viral RNA
4. Interaction of APOBEC3G with HIV-1 Gag proteins. Only the latter two mechanisms have been extensively confirmed.
Fig.2
Vif also has an important role in APOBEC3G's turnover. It impairs the translation of hA3G mRNA, probably through an mRNA-binding mechanism and in particular binding to hA3G protein recruits an E3 ubiquitin ligase that mediates the polyubiquitylation of hA3G and its degradation by proteasome.
REGULATION
The Apo3G dimer is predicted to form via head-to-head interactions with residues on the CD1 domains (Fig. 1). Mutagenesis data identifies Apo3G residues YYFW124-127 residing on Loop 7 of the CD1 domain as important for dimerization. In the CD2 domain, the corresponding residues on Loop 7 are important for substrate specificity; however, in the proposed Apo3G dimer, the Loop 7 CD1 residues are buried. In this type of Apo3G dimerization, the CD1 active sites would not be accessible to ssDNA substrates, which may explain why the CD1 domains are inactive. Other models proposed that Apo3G forms dimers via interactions of the CD2 domain. Fret and co-immunoprecipitation experiments show that the truncated Apo3G-CD1 proteins do not self associate, while the truncated Apo3G-CD2 proteins do. These results remain controversial as two other reports demonstrate that Apo3G-CD2 protein exists as a monomer. However, it is possible that Apo3G may form dimers and higher order oligomers through head-to-head interactions of both, the CD1 and CD2 domains. Additionally, Apo3G oligomerization state is influenced by salt concentrations and the presence of DNA or RNA.
A model for oligomeric regulation of APOBEC3G cytosine deaminase-dependent restriction of HIV. 2008
DISEASES RELEVANCE
APOBEC3G is expressed within the non-permissive cells and is a key inhibitory factor of HIV-1 replication and infectivity. However, Vif counteracts this antiretroviral factor, enabling production of viable and infective HIV-1 virions in the presence of APOBEC3G activity. In particular, Vif prevents incorporation of APOBEC3G into HIV-1 virions and promotes destruction of the enzyme in a manner independent of all other HIV-1 proteins.
While APOBEC3G has typically been studied as a vital protein exhibiting potent antiviral effects on HIV-1, recent studies have elucidated the potential of APOBEC3G-mediated mutation to help to facilitate the propagation HIV-1. The number of deaminations in the preferred regions varies from one to many, possibly dependent on the time of exposure to APOBEC3G.
APOBEC3G contributes to HIV-1 variation through sublethal mutagenesis. 2010
Additionally, it has been shown that there is a dose response between intracellular APOBEC3G concentration and degree of viral hypermutation.
Turning up the volume on mutational pressure: is more of a good thing always better? (A case study of HIV-1 Vif and APOBEC3). 2008
Some HIV-1 proviruses with APOBEC3G-mediated mutation have been shown to thrive because they carry too few mutations at APOBEC3G hotspots or because recombination between a lethally APOBEC3G-restricted provirus and a viable provirus has occurred. Such sublethal mutagenesis contributes to greater genetic diversity among the HIV-1 virus population, demonstrating the potential for APOBEC3G to enhance HIV-1’s ability to adapt and propagate.
