Comparative lipidomics analysis of HIV-1 particles and their producer cell membrane in different cell lines

Abstract

SummaryHuman immunodeficiency virus type 1 (HIV-1) is a retrovirus that obtains its lipid envelope by budding through the plasma membrane of infected host cells. Various studies indicated that the HIV-1 membrane differs from the producer cell plasma membrane suggesting virus budding from preexisting subdomains or virus-mediated induction of a specialized budding membrane. To perform a comparative lipidomics analysis by quantitative mass spectrometry, we first evaluated two independent methods to isolate the cellular plasma membrane. Subsequent lipid analysis of plasma membranes and HIV-1 purified from two different cell lines revealed a significantly different lipid composition of the viral membrane compared with the host cell plasma membrane, independent of the cell type investigated. Virus particles were significantly enriched in phosphatidylserine, sphingomyelin, hexosylceramide and saturated phosphatidylcholine species when compared with the host cell plasma membrane of the producer cells; they showed reduced levels of unsaturated phosphatidylcholine species, phosphatidylethanolamine and phosphatidylinositol. Cell type-specific differences in the lipid composition of HIV-1 and donor plasma membranes were observed for plasmalogenphosphatidylethanolamine and phosphatidylglycerol, which were strongly enriched only in HIV-1 derived from MT-4 cells. MT-4 cell-derived HIV-1 also contained dihydrosphingomyelin as reported previously, but this lipid class was also enriched in the host cell membrane. Taken together, these data strongly support the hypothesis that HIV-1 selects a specific lipid environment for its morphogenesis.

Analysis of lipid-composition changes in plasma membrane microdomains

“…The yield of SCM lipids was ‫ف‬ 5% that of WC lipids ( Fig. 1B ), a reasonable value in view of the proportion of plasma membranes to WC membranes ( 31,37 ). These results showed that the SCMs obtained in this study comprised a plasma membrane-rich fraction, while it also may have contained some intracellular components tightly associated with the plasma membrane via cytoskeletal proteins.…”

“…After mechanical cell lysis, the SCMs can be separated by centrifugation, due to their high density. However, purities and yields of the membranes can vary in different laboratories because of the different experimental conditions used, including variations in buffer systems (26)(27)(28)(29)(30)(31)(32)(33)(34). In addition, a relatively large number of cells were needed in previous plasma membrane lipidomics studies using this method ( 30,31 ).…”

“…1). The recovery rate of the detergent-resistant membrane lipid was over 10% of the WC lipid, suggesting that the drFF also contained membranes other than plasma membrane microdomains ( 31,37 ). The values were relatively variable between independent experiments.…”

“…The cationic colloidal silica beads method was used for preparing plasma membranes (26)(27)(28)(29)(30)(31)(32)(33)(34). Briefl y, this step was performed on ice using ice-cold buffers.…”

Mcp1 and Mcp2, two novel proteins involved in mitochondrial lipid homeostasis

“…Mass spectrometry analysis was performed in positive ion mode on a triple quadrupole-linear ion trap hybrid mass spectrometer (QTRAP 5500, AB Siex), except for analysis of cardiolipin and ergosterol, which was done in negative ion mode on a quadrupole time-of-flight mass spectrometer (QStar Elite, AB Siex). Lipid extractions (total lipid amount of 1.5-2.5 nmol) were performed in the presence of internal lipid standards [PC, PE, PS with species 14:1/14:20:1/20:1 and 22:1/22:1), PI (37:4), CL 56:0] (Lorizate et al, 2013). Dried lipids were dissolved in 10 mM ammonium acetate.…”