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That assembly and release of HIV-1 particles takes place at the plasma membrane has been established for some time. It has also been suspected that the process additionally requires lipid rafts — plasma membrane domains that serve as docking sites for some proteins involved in signal transduction. In 20 November Proceedings of the National Academy of Sciences, Akira Ono and Eric Freed at the National Institute of Allergy and Infectious Diseases (NIAID) Laboratory of Molecular Microbiology, have shown not only that the viral Gag protein, which promotes particle production, associates with lipid rafts, but also that the protein's N terminus and the Gag-Gag interaction domains are crucial for the association (Proc Natl Acad Sci USA 2001, 98:13925-13930).This has increased speculation that cholesterol-depleting drugs might suppress HIV replication in people. Rafts are rich in cholesterol as well as sphingolipids. James Hildreth, Associate Professor of Pharmacology and Molecular Science, along with his colleagues at Johns Hopkins University in Baltimore, reported in July AIDS Research and Human Retroviruses that draining cholesterol from the cell membrane can completely block HIV transmission. Based on preliminary success in a murine model, Hildreth suggested that sexually transmitted HIV might be prevented with a kind of chemical condom, a topical cream containing an otherwise-harmless compound that attacks cholesterol.Ono and Freed observed that depleting cellular cholesterol reduces HIV-1 particle production and that disrupting lipid rafts cripples the virus's infectivity. But Freed urged caution in making the "somewhat risky" leap from these lab findings to clinical applications. "I would not expect cholesterol-lowering drugs to prevent HIV infection. It remains to be tested, however, whether these drugs might reduce viral loads in infected individuals. If so, they might show some clinical benefits when combined with other antiviral agents," he told BioMed Central."Although I personally am not very convinced that these drugs may be sufficient to stop viral progression, it is possible that they may help in combinations with current treatments to delay the propagation of the virus in the body," agreed Santos Mañes from the Department of Immunology and Oncology of the Centro Nacional de Biotecnología at the Universidad Autonoma de Madrid.One way to establish that would be to look at HIV-1 progression in infected patients who are on cholesterol-lowering drugs for other diseases, according to Robert Blumenthal, of the Laboratory of Experimental and Computational Biology at the National Cancer Institute. Blumenthal — whose research focuses on rafts in viral entry — co-organized a meeting on the topic in Spring 2000.Johnson Mak and colleagues at the Macfarlane Burnet Centre for Medical Research, Fairfield, Victoria, Australia, reviewed research on the relationship between lipid rafts and HIV-1 in October 2001 Journal of Clinical Virology. He said that unpublished data from his lab demonstrated that removal of cholesterol from the virion completely destroys HIV-1, causing it to collapse and become non-infectious. "Targeting cholesterol to reduce HIV-1 infectivity clearly is an approach that we are pursuing," he told BioMed Central. But, he warned that the complexity of cholesterol metabolism and balance in the host is difficult to manage.Mak regarded it as unlikely that currently available cholesterol–lowering compounds can suppress HIV-1 replication effectively — although he added a caveat: "never say never." The key is continued study of the relationship between lipid and HIV-1 via basic research. "This will help us to better understand this relationship and then to design more viral-specific targets to interfere with the lipid/HIV-1 interplay that does not affect the host. An exciting start, but clearly not there yet."Another intriguing possibility is that work on lipid rafts may reveal much about other diseases. A variety of enveloped viruses that are human pathogens — among them influenza and measles — appear to replicate in a raft-dependent manner. So strategies that would combat HIV infection might also show some effects in these viral systems.Mak conceded that the exact role of lipid rafts in infectious disease is debatable. But he also thinks speculation is warranted. In the October 2001 review, his team list more than a dozen pathogens already known to require either cholesterol or lipid rafts to cause disease or establish infection. In addition to several viruses, the list includes mycobacteria, the beta-amyloid of Alzheimer's disease, some forms of Escherichia coli, and the scrapie isoform of the prion protein. "I would imagine that the list will continue to grow as we look closer. After all, lipid rafts are key players in signaling events, and pathogens might have evolved to attack strategic key points in the cell to establish productive infection."In order to establish that the Gag protein is physiologically relevant in virus assembly, Ono and Freed used two classes of compounds to disrupt rafts: a cyclic sugar compound known as MβCD, which can extract cholesterol from plasma membrane, and the clinically ubiquitous statins, which block cholesterol biosynthesis. "But despite these differing modes of action, both compounds have similar effects in inhibiting virus production," Freed explained.They used a variety of controls to make sure that the defect in virus production could not simply be a result of global cellular toxicity. These included several different Gag mutants and other viruses. In contrast to wild-type Gag, release of Gag mutants that lack a late domain that is important for the final budding step was not reduced by cholesterol depletion. "These controls indicate that reduction in virus release that we observe with wild-type Gag is not just the result of local cellular toxicity, but is a specific process for affecting wild-type HIV-1 Gag", Freed notedBy understanding how Gag goes out from the host cell to form virus particles, researchers can also get an idea of how the virus might enter the cell, Mak said. He believes it is possible that the exit and entry of virus from their host or target cells, respectively, are mirror-image events, at least in terms of the role of lipid in virion assembly for release and virion uncoating during entry. "By understanding their interplay, I am confident that we can design better drugs against HIV-1," he declared.What's next for the researchers? Freed said his lab will continue to investigate the role of rafts in HIV replication from the perspective of basic virology and cell biology. "Of special interest to us is the possibility that the targeting of assembly to rafts might be particularly significant in facilitating cell-cell virus transmission," he reported.Mañes argued that in order to really understand HIV-1 budding, researchers need to figure out which specific steps in viral production require rafts. "From this and other related papers I have the impression that rafts are required for different steps in viral budding but not for all," he noted. Mañes thinks identification of such steps could help imagine new ways to affect HIV budding — and hence HIV-1's ability to infect.

HIV entry and membrane lipid rafts

Cholesterol lowering drugs could be used to disrupt the cell entry mechanism of many intracellular pathogens.

Dec 3, 2001

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