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The seasonal variation of CuCl2-mediated low density lipoprotein (LDL) oxidation (10 microM Cu2+, lag phase, rate of oxidation and maximum absorbance at 234 nm) were measured in 43 men and women on 4-6 occasions (mean 5.7 +/- 0.5) over a 12-month period. The lag phase averaged 52.7 +/- 0.6 min and did not differ by gender. Lag phase and rate of the rapid propagation phase of LDL oxidation showed a sinusoidal pattern over the year (increased and reduced oxidative susceptibility during January and June-July, respectively; both p < 0.001). Changes in plasma alpha-tocopherol, ascorbic acid, lycopene or beta-carotene concentrations did not explain seasonal differences in oxidative susceptibility of LDL in vitro. Nor did plasma lipid content of linoleic acid, the main substrate of lipid peroxidation, vary. However, the amount of hydroperoxy- plus hydroxy-fatty acids in plasma lipids varied according to season (p < 0.024) and was related to the lag phase (r = -0.26, p < 0.001). Seasonal variation in oxidative susceptibility was not significant after adjusting for hydroperoxy- plus hydroxy-fatty acids (p = 0.506). Isolated LDL is more vulnerable to Cu2+-induced lipid peroxidation during the winter and this may be due to the higher amount of oxidised lipids during that period.

Heterologous expression of HIV-1 Gag in a variety of host cells results in its packaging into virus-like particles (VLPs) that are subsequently released into the extracellular milieu. This phenomenon represents a useful tool for probing cellular factors required for viral budding and has contributed to the discovery of roles for ubiquitin ligases and the endosomal sorting complexes required for transport (ESCRTs) in viral budding. These factors are highly conserved throughout eukaryotes and have been studied extensively in the yeast Saccharomyces cerevisiae, a model eukaryote previously utilized as a host for the production of VLPs. We used heterologous expression of HIV Gag in yeast spheroplasts to examine the role of ESCRTs and associated factors (Rsp5, a HECT ubiquitin ligase of the Nedd4 family; Bro1, a homolog of Alix; and Vps4, the AAA-ATPase required for ESCRT function in all contexts/organisms investigated) in the generation of VLPs. Our data reveal: 1) characterized Gag-ESCRT interaction motifs (late domains) are not required for VLP budding, 2) loss of function alleles of the essential HECT ubiquitin ligase Rsp5 do not display defects in VLP formation, and 3) ESCRT function is not required for VLP formation from spheroplasts. These results suggest that the egress of HIV Gag from yeast cells is distinct from the most commonly described mode of exit from mammalian cells, instead mimicking ESCRT-independent VLP formation observed in a subset of mammalian cells. As such, budding of Gag from yeast cells appears to represent ESCRT-independent budding relevant to viral replication in at least some situations. Thus the myriad of genetic and biochemical tools available in the yeast system may be of utility in the study of this aspect of viral budding.