Human hepatic lipase. Cloned cDNA sequence, restriction fragment length polymorphisms, chromosomal localization, and evolutionary relationships with lipoprotein lipase and pancreatic lipase.

The addition and endoplasmic reticulum (ER) glucosidase processing of N-linked glycans is essential for the secretion of rat hepatic lipase (HL). Human HL is distinct from rat HL by the presence of four as opposed to two N-linked carbohydrate side chains. We examined the role of N-linked glycosylation and calnexin interaction in human HL secretion from Chinese hamster ovary (CHO) cells stably expressing a human HL cDNA. Steady-state and pulse-chase labeling experiments established that human HL was synthesized as an ER-associated precursor containing high mannose N-linked glycans. Secreted HL had a molecular mass of ϳ 65 kDa and contained mature N-linked sugars. Inhibition of N-linked glycosylation with tunicamycin (TM) prevented secretion of HL enzyme activity and protein mass. In contrast, incubation of cells with the ER glucosidase inhibitor, castanospermine (CST), decreased human HL protein secretion by 60%, but allowed 40% of fully active HL to be secreted. HL protein mass and enzyme activity were also recovered from the media of a CHO-derivative cell line genetically deficient in ER glucosidase I activity (Lec23) that was transiently transfected with a human HL cDNA. Co-immunoprecipitation experiments demonstrated that newly synthesized human HL bound to the lectin-like ER chaperone, calnexin, and that this interaction was inhibited by TM and CST.These results suggest that under normal conditions calnexin may increase the efficiency of HL export from the ER. Whereas a significant proportion of human HL can attain activity and become secreted in the absence of glucose trimming and calnexin association, these interrelated processes are nevertheless essential for the expression of full HL activity. -Boedeker, J. C., M. Doolittle, S. Santamarina-Fojo, and A. L. White. Role of N-linked carbohydrate processing and calnexin in human hepatic lipase secretion.

Section: Synthesis and Secretion Of Human Hl In Stably Transfected Cho Cell Culturessupporting

confidence: 68%

mentioning

confidence: 91%

Role of N-linked carbohydrate processing and calnexin in human hepatic lipase secretion

Boedeker

Doolittle

Santamarina-Fojo

et al. 1999

“…Fuki et al (46) described an analogous mechanism in vitro. In those studies, lipoproteins enriched in lipoprotein lipase, which is structurally and functionally similar to HL (31,47,48), were internalized through a process mediated by the proteoglycan syndecan-1.…”

Section: Discussionmentioning

confidence: 99%

Hepatic lipase overexpression lowers remnant and LDL levels by a noncatalytic mechanism in LDL receptor-deficient mice

Dichek

Johnson²,

Akeefe

et al. 2001

To address the role of the noncatalytic ligand function of hepatic lipase (HL) in low density lipoprotein (LDL) receptor-mediated lipoprotein metabolism, we characterized transgenic mice lacking the LDL receptor (LDLR) that express either catalytically active ( Ldlr ؊ / ؊ HL ) or inactive ( Ldlr ؊ / ؊ HL S145G ) human HL on both chow and high fat diets and compared them with nontransgenic Ldlr ؊ / ؊ mice. In mice fed a chow diet, apolipoprotein (apo)B-containing lipoprotein levels were 40-60% lower in Ldlr ؊ / ؊ HL and Ldlr ؊ / ؊ HL S145G mice than in Ldlr ؊ / ؊ mice. This decrease was mainly reflected by decreased apoB-48 levels in the Ldlr ؊ / ؊ HL mice and by decreased apoB-100 levels in Ldlr ؊ / ؊ HL S145G mice. These findings indicate that HL can reduce apoB-100-containing lipoproteins through a noncatalytic ligand activity that is independent of the LDLR. Cholesterol enrichment of the apoB-containing lipoproteins induced by feeding Ldlr ؊ / ؊ HL and Ldlr ؊ / ؊ HL S145G mice a cholesterolenriched high fat (Western) diet resulted in parallel decreases in both apoB-100 and apoB-48 levels, indicating that HL is particularly efficient at reducing cholesterol-enriched apoB-containing lipoproteins through both catalytic and noncatalytic mechanisms. These data suggest that the noncatalytic function of HL provides an alternate clearance pathway for apoB-100-and apoB-48-containing lipoproteins that is independent of the LDLR and that contributes to the clearance of high density lipoproteins.

“…Hepatic lipase (HL) is a 477-amino acid glycoprotein that plays a pivotal role in lipoprotein metabolism (1). The human HL gene spans more than 30 kb, is composed of nine exons and eight introns (2,3), and is located on chromosome 15q21 (4). The majority of HL is synthesized and secreted by the liver and is bound to heparan sulfate proteoglycans on the surfaces of sinusoidal endothelial cells and external surfaces of microvilli of parenchymal cells in the space of Disse (5,6).…”

mentioning

confidence: 99%

In vivo evidence of a role for hepatic lipase in human apoB-containing lipoprotein metabolism, independent of its lipolytic activity

Zambón

Deeb

Bensadoun

et al. 2000

Hepatic lipase (HL) is a key player in lipoprotein metabolism by modulating, through its lipolytic activity, the triglyceride (TG) and phospholipid content of apolipoprotein B (apoB)-containing lipoproteins and of high density lipoproteins (HDL), thereby affecting their size and density. A new and separate role has been suggested for HL in cellular lipoprotein metabolism, in which it serves as a ligand promoting cellular uptake of apoB-containing remnant lipoproteins and HDL. We tested the hypothesis that HL has both a lipolytic and a nonlipolytic role in human lipoprotein metabolism, by measuring lipid plasma concentrations, lipoprotein density distribution by density gradient ultracentrifugation, and lipoprotein composition, in three subjects with HL deficiency: two of the patients (S-1 and S-3) were characterized as having neither plasma HL activity nor detectable HL protein; the third subject (S-2) had no plasma HL activity but a detectable amount (35.5 ng/ml) of HL protein. All HL-deficient subjects showed a severalfold increase in lipoprotein TG content across the lipoprotein density spectrum [very low density lipoprotein (VLDL), intermediate density lipoprotein (IDL), low density lipoprotein (LDL), and HDL] as compared with control subjects. They also had remarkably more buoyant LDL particles (LDL-R f ‫؍‬ 0.342-0.394) as compared with the control subjects (LDL-R f ‫؍‬ 0.303). Subjects S-1 and S-3 (no HL activity or protein) presented with a distinct increase in cholesterol and apoB levels in the IDL and VLDL density range as compared with patient S-2, with detectable HL protein, and the control subjects. This study provides evidence in humans that HL indeed plays an important role in lipoprotein metabolism independent of its enzymatic activity: in particular, inactive HL protein appears to affect VLDL and IDL particle concentration, whereas HL enzymatic activity seems to influence VLDL-, IDL-, LDL-, and HDL-TG content and their physical properties. -Zambon, A., S. S. Deeb, A. Bensadoun, K. E. Foster, and J. D. Brunzell. In vivo evidence of a role for hepatic lipase in human apoB-containing lipoprotein metabolism, independent of its lipolytic activity.