Abstract
Endothelial lipase (EL) is a new member of the triglyceride lipase gene family, which includes lipoprotein lipase (LpL) and hepatic lipase (HL). Enzymatic activity of EL has been studied before. Here we characterized the ability of EL to bridge lipoproteins to the cell surface. Expression of EL in wild-type Chinese hamster ovary (CHO)-K1 but not in heparan sulfate proteoglycan (HSPG)-deficient CHO-677 cells resulted in 3-4.4-fold increases of 125I-low density lipoprotein (LDL) and 125I-high density lipoprotein 3 binding (HDL3). Inhibition of proteoglycan sulfation by sodium chlorate or incubation of cells with labeled lipoproteins in the presence of heparin (100 microg/ml) abolished bridging effects of EL. An enzymatically inactive EL, EL-S149A, was equally effective in facilitating lipoprotein bridging as native EL. Processing of LDL and HDL differed notably after initial binding via EL to the cell surface. More than 90% of the surface-bound 125I-LDL was destined for internalization and degradation, whereas about 70% of the surface-bound 125I-HDL3 was released back into the medium. These differences were significantly attenuated after HDL clustering was promoted using antibody against apolipoprotein A-I. At equal protein concentration of added lipoproteins the ratio of HDL3 to VLDL bridging via EL was 0.092 compared with 0.174 via HL and 0.002 via LpL. In summary, EL mediates binding and uptake of plasma lipoproteins via a process that is independent of its enzymatic activity, requires cellular heparan sulfate proteoglycans, and is regulated by ligand clustering.
Highlights
Effects of Endothelial lipase (EL) Expression on Binding of 125I-low density lipoprotein (LDL) and 125IHDL3—We examined the effects of EL on the binding of 125I-labeled LDL (125ILDL) and 125I-high density lipoprotein 3 binding (HDL3) in Chinese hamster ovary (CHO)-K1 cells
125ILDL or 125I-HDL3 were added to the cells at equal protein concentrations, about 2.3 times more 125I-LDL protein was bound to the cells compared with 125I-HDL3 via the EL-mediated process (114.3 Ϯ 7.6 versus 49.4 Ϯ 2.3 ng/mg of cell protein, respectively) indicating that a higher percentage of LDL particles was bound to the cells compared with HDL
The results presented in this study demonstrate that EL, a recently discovered member of the triglyceride lipase family, is able to mediate efficient binding and cellular metabolism of both apoB- and apoA-I-containing lipoproteins
Summary
LpL, lipoprotein lipase; anti-apoA-I, hamster ovary; EL, endothelial lipase; HL, hepatic lipase; HS, heparan polyclonal goat antibody against human apolipoprotein A-I; AdGFP, sulfate; HSPG, heparan sulfate proteoglycan; HDL, high density liadenovirus encoding GFP; adEL, adenovirus encoding endothelial poprotein; LDL, low-density lipoprotein; VLDL, very low density lilipase; adEL-S149A, adenovirus encoding catalytically inactive endo- poprotein; TG, triglyceride; GFP, green fluorescent protein; DiI, thelial lipase; AdHL, adenovirus encoding hepatic lipase; adLpL, ade- 1,1Ј-dioctadecyl-3,3,3Ј,3Ј-tetramethylindocarbocyanine. We found that compared with LpL and HL, EL has distinct preferences in bridging individual classes of lipoproteins
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