All modern viral pandemics in humans have been the result of cross-species transmissions of virus from another host into the human population.  These cross-species transmissions, called zoonoses, are the limited by the ability of virus to adapt to use proteins of its new host, and by the ability of the virus to escape from human antiviral proteins.  These defenses, called restriction factors, consist of an array of proteins that interact with and block the activity of viral pathways necessary for infection, replication, and transmission. 

The natural history of the transmission of what the human immunodeficiency virus-1 (HIV1) from primates to humans is a tale of just such an adaptation to new hosts.  A simian immunodeficiency virus (SIV) of monkeys was transmitted to chimpanzees in the ancient past.   This virus of chimpanzees, called SIVcpz, was subsequently transmitted to humans several times in the past century and is the origin of the HIV-1 pandemic.   In order to productively infect chimpanzees and then humans, the precursor of HIV needed to overcome existing host defense mechanisms such as the apolipoprotein B mRNA editing enzyme catalytic polypeptide-like 3G, APOBEC3G or A3G enzyme. The A3G deaminase converts cytidine residues within the single-stranded DNA HIV replication intermediates into uridines, effectively blocking viral replication.  To combat this activity, lentiviruses including HIV have evolved a viral infectivity factor or Vif that binds to A3G and targets it for proteosomal degradation.  On a molecular level, the specificity of the interaction between Vif and primate A3G is determined by individual amino acids that for the protein/protein interface.  For A3G, the critical residues encode aspartic acid at position 128 and 130 (D/D) in hominids including humans and chimpanzees.  Monkeys, on the other hand, such as macaques and guenons for example, have lysine 128 and asparagine 130 (K/N) and glutamic acid 128 and alanine 130 (E/A), respectively.  These differences confer host specific A3G recognition of lentiviral Vif, and Vif must adapt to the new interface for the virus to transfer between hosts.  In a previous study the Emerman lab in collaboration with the Gross lab at UCSF found that the Vif gene from an SIV from a monkey needed to adapt to to the D/D at positions 128/130 in chimpanzee A3G and did so by changes in one particular loop of Vif.   In this study, postdoctoral fellow Nicholas Chesarino in Michael Emerman’s Human Biology Division laboratory set out to determine if this adaptation of Vif to a new interface resulted in a switch in specificity to the hominid version of A3G, or whether Vif retained its original specificity and then broadened its specificity to new interfaces.     The results were recently posted on the preprint server biorxiv.

To test the Vif antagonism of all potential amino acid combinations at A3G positions 128 and 130, a library containing degenerate NNS (N is any base, S is cytosine or guanine) codons was constructed.  Use of the NNS codon allows for all natural amino acids but only one of the three possible stop codons, thus limiting expression of truncated mutants.  The resulting library of mutant A3G proteins was expressed and tested in the presence or absence of Vif for HIV restriction.  175 unique A3G clones were tested yielding distinct categories of mutant proteins.  First were A3G variants that restricted HIV infection in the absence of Vif, essentially possessing wild type anti-viral activity.  As expected, proteins containing internal stop codons were inactive, as were several full-length proteins.  Of the single amino acid variants at positions 128 and 130, 91% retained antiviral activity while only 30% of double mutants were still active.  Based on these results, a global map of A3G mutations at positions 128 and 130 was constructed.   The results showed that that the specificity of the Vif gene of HIV-1 was broadened after its transfer from monkeys to chimps to humans.   Thus, cross-species transmission of the lentiviruses tends to broaden their specificity rather than narrowing it to the new host.

While HIV-1 Vif gained a broad ability to recognize divergence at the critical 128/130 interface, some mutations in A3G allowed it to escape HIV-1 Vif antagonism.  Synthetic DNA libraries can be designed to encode any amino acid at any position, but evolution is more constrained.  Changes in three nucleotide codons are most likely to occur one nucleotide at a time and not all amino acid transitions are possible through a single nucleotide change.  An additional constraint is that intermediate steps involving amino acids other than first and last need to retain functionality of the protein.  Examination of the codons that allowed resistance to Vif antagonism versus those that are attainable by single nucleotide mutations, showed that an evolutionarily accessible path to resistance by the hominid versions of A3G.  Thus, while hominids are protected from infection from most SIVs because their A3G is resistant to antagonism by the SIV Vif gene encoded by most monkey SIVs, hominids such as humans have also reached a dead-end in their ability to further evolve to resist modern SIVs such as SIVcpz in chimpanzees and HIV-1 in humans.   

Chesarino M, Emerman M. HIV-1 Vif gained breadth in APOBEC3G specificity afer cross-species transmission of its precursors. biorXiv. 2021. doi: https://doi.org/10.1101/2021.08.23.457421

This research was supported by grants from the National Institute of Allergies and Infectious Diseases.

Fred Hutch/UW Cancer Consortium member Michael Emerman contributed to this study.