Caveolin-1β promotes the production of active human microsomal glutathione S-transferase in induced intracellular vesicles inSpodoptera frugiperda21 insect cells

Abstract

The heterologous expression in Spodoptera frugiperda 21 (Sf21) insect cells of the β isoform of canine caveolin-1 (caveolin-1β), using a baculovirus-based vector, resulted in intracellular vesicles enriched in caveolin-1β. We investigated whether these vesicles could act as membrane reservoirs, and promote the production of an active membrane protein (MP) when co-expressed with caveolin-1β. We chose hMGST1 (human microsomal glutathione S-transferase 1) as the co-expressed MP. It belongs to the membrane-associated proteins in eicosanoid and glutathione metabolism (MAPEG) family of integral MPs, and, as a phase II detoxification enzyme, it catalyzes glutathione conjugation of lipophilic drugs present in the lipid membranes. In addition to its pharmaceutical interest, its GST activity can be conveniently measured. The expression of both MPs were followed by Western blots and membrane fractionation on density gradient, and their cell localization by immunolabeling and transmission electron microscopy. We showed that caveolin-1β kept its capacity to induce intracellular vesicles in the host when co-expressed with hMGST1, and that hMGST1 is in part addressed to these vesicles. Remarkably, a fourfold increase in the amount of active hMGST1 was found in the most enriched membrane fraction, along with an increase of its specific activity by 60% when it was co-expressed with caveolin-1β. Thus, heterologously expressed caveolin-1β was able to induce cytoplasmic vesicles in which a co-expressed exogenous MP is diverted and sequestered, providing a favorable environment for this cargo.

Keywords: Canine caveolin-1β; Heterologous co-expression; Human MGST1; Membrane protein; Membrane remodeling; Sf21 insect cells.