259 In Vitro HIV-1 Infection Increases Intracellular Cholesterol A.B. van 't Wout*, D.M. Cunningham, H. He, G.K. Geiss, J.I. Mullins Univ of Washington, Seattle
Background: Recent studies by several laboratories have highlighted the importance of cholesterol in the HIV-1 life cycle. Removing cholesterol from virions or inhibition of cholesterol biosynthesis with lovastatin inhibits HIV-1 production in vitro. This inhibition is thought to be mediated through the cholesterol-rich domains on the cell surface (lipid rafts) through which HIV-1 emerges during viral maturation. Our microarray studies of gene expression in CD4+ T-cell lines infected with HIV-1 in vitro indicated specific up-regulation of genes involved in sterol biosynthesis. Expression of the LDL receptor, responsible for cholesterol uptake, was also increased.
Methods: High-density cDNA microarrays were used to measure changes in gene expression. VSV-HIV pseudotypes and chemokine receptor stimulation were tested to distinguish between envelope-dependent and -independent effects. Cells exposed to heat shock, interferon, or influenza A infection were analyzed to determine the specificity of the HIV-1 induced changes. To determine the impact of transcriptional activation on the levels of sterol biosynthesis enzymes, cells were labeled with 14C mevalonic acid, an upstream precursor to the sterol biosynthesis pathway, and 14C incorporation in downstream products was assessed.
Results: To determine the HIV gene product(s) responsible for the induction of the cholesterol pathway, we studied envelope-deleted virus variants and signaling through CCR5 or CXCR4. Results suggested that the virus envelope was responsible for part of the induction, but that other viral gene products must also be involved. The impact of individual gene products is under study. The induction was not part of a general stress response since it was not detected in response to heat shock, interferon treatment, or infection with Influenza A. Consistent with the microarray results, the rate of 14C mevalonic acid incorporation was increased 2.5-fold in infected CEM cells, and 7.4 fold in SupT1 cells.
Conclusions: Our studies indicate that HIV-1 infection promotes the accumulation of intracellular cholesterol by increasing both cholesterol production and uptake, thereby enhancing the release of infectious viral particles. Hence, cholesterol production and uptake should be evaluated as a target for anti-HIV therapy.
