BioWorld International Correspondent
LONDON - A family of enzymes that makes proteins ready for processing by the cell's waste-disposal machinery could be an ideal drug target for therapies against HIV.
Earlier this year, several studies showed that the Vif protein of HIV-1 disabled a cellular protein called APOBEC3G, which normally has the function of protecting cells against viral infection.
Now, two studies in Nature Medicine (published online Oct. 5, 2003) show that Vif both binds to APOBEC3G and processes it so that it can be degraded by the proteasome, a cellular organelle that breaks down proteins that are no longer needed.
Michael Malim, who led one of the teams that had its work published in Nature Medicine, told BioWorld International: "If you could interfere with Vif's ability to induce degradation of APOBEC3G, maybe levels of this cellular protein would remain sufficiently high enough for it to become incorporated into viral particles, and [thereby] block infection."
Malim, head of infectious diseases at King's College London, and his colleagues identified the gene encoding APOBEC3G more than a year ago. Earlier this year, several other groups described how APOBEC3G has an antiretroviral effect by bringing about mutations in single-stranded DNA made by HIV.
The studies showed that APOBEC3G changes cytidine bases to uridine in single-stranded DNA of viral origin, which has the effect of changing guanine bases to adenine in the opposing DNA strand. Those mutations cause stop codons in viral proteins, thereby blocking virion production.
The pair of Nature Medicine papers added more detail to how Vif combats the protective function of APOBEC3G. Malim - with colleagues Ann Sheehy, also of King's College London, and Nathan Gaddis, of the University of Pennsylvania School of Medicine in Philadelphia - reported his findings in a paper titled "The antiretroviral enzyme APOBEC3G is degraded by the proteasome in response to HIV-1 Vif."
David Kabat, at the Oregon Health and Sciences University in Portland, and colleagues described their study in a paper titled "HIV-1 Vif protein binds the editing enzyme APOBEC3G and induces its degradation."
The King's group transfected T cells with an expression vector containing either wild-type HIV-1 DNA or HIV-1 DNA lacking the gene encoding Vif. After 24 hours, the protein contents of virions produced by these cells were analyzed. The team found that virions produced by cells transfected by wild-type HIV-1 DNA contained virtually no APOBEC3G, while APOBEC3G was present in those that lacked Vif.
Further experiments showed that the presence of Vif caused a substantial fall in expression of APOBEC3G, but that Vif did not affect levels of messenger RNA for APOBEC3G. The team went on to show that, in general, the presence of Vif reduced the half-life of APOBEC3G protein by about 50 percent, from about seven hours to 3.5 hours.
To investigate whether the cells were getting rid of APOBEC3G via the proteasome, the next experiment involved adding substances that inhibited that pathway. When the inhibitors were present, levels of APOBEC3G returned to normal. The researchers wrote, "The reduction in APOBEC3G stability in cells containing Vif is attributable to the induction of proteasome-mediated degradation."
Malim told BioWorld International: "We would like to know what other cellular proteins are involved in mediating this degradative effect. If a protein is to be targeted to the proteasome, it must first be modified by small cellular proteins called ubiquitin. Appending ubiquitin requires enzymes which are presumably supplied by the host cell - but the ubiquitin ligases are a very large family."
Kabat and his colleagues concluded that their results are "encouraging from a drug development perspective." The team found that one region of Vif binds APOBEC3G, while another region - which is conserved - is involved in directing APOBEC3G for degradation by the proteasome. Drugs targeted at that evolutionarily conserved site might, the researchers suggest, be resistant to viral escape mutations. They also suggested exploring drugs that inhibit Vif's binding to APOBEC3G.
Kabat and his team concluded in Nature Medicine, "These considerations suggest that drug selection and evaluation strategies focused on the binding and degradation processes described here may be worthwhile."