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Molecular characterization of quail apolipoprotein very-low-density lipoprotein II: disulphide-bond-mediated dimerization is not essential for inhibition of lipoprotein lipase.

ABSTRACT: As part of the avian reproductive effort, large quantities of triglyceride-rich very-low-density lipoprotein (VLDL) particles are transported by receptor-mediated endocytosis into the female germ cells. Although the oocytes are surrounded by a layer of granulosa cells harbouring high levels of active lipoprotein lipase, non-lipolysed VLDL is transported into the yolk. This is because VLDL particles from laying chickens are protected from lipolysis by apolipoprotein (apo)-VLDL-II, a potent dimeric lipoprotein lipase inhibitor [Schneider, Carroll, Severson and Nimpf (1990) J. Lipid Res. 31, 507-513]. To determine whether this protection depends on dimer formation and constitutes a general mechanism to ensure high levels of yolk triglycerides for embryonic utilization in birds, we have now molecularly characterized apo-VLDL-II in the Japanese quail, a frequently used avian species. Quail apo-VLDL-II shows 72% amino acid identity with the chicken protein, with most replacements being in the C-terminal region. Importantly, quail apo-VLDL-II lacks the single cysteine residue present eight residues from the C-terminus of chicken apo-VLDL-II, which is responsible for dimerization of the chicken lipoprotein lipase inhibitor. Nevertheless, monomeric quail and dimeric chicken apo-VLDL-II display, on a molar basis, identical inhibitory effects on lipoprotein lipase, underscoring the biological importance of their function. Furthermore secondary structure prediction of the 3'-untranslated region of the quail message supports a role for loop structures in the strictly oestrogen-dependent production of the lipoprotein lipase inhibitors. Our findings shed new light on the essential role of this small, hormonally regulated, protein in avian reproduction.

SUBMITTER: MacLachlan I

PROVIDER: S-EPMC1217528 | biostudies-other | 1996 Jul

REPOSITORIES: biostudies-other

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Project description:The function of apolipoprotein (apo) C1 in vivo is not clearly defined. Because transgenic mice overexpressing human apoC1 show elevated triacylglycerol (TG) levels [Simonet, Bucay, Pitas, Lauer and Taylor (1991) J. Biol. Chem. 266, 8651-8654], an as yet unknown role for apoC1 in TG metabolism has been suggested. Here we investigated directly the effect of the complete absence of apoC1 on very-low-density lipoprotein (VLDL)-TG lipolysis, clearance and production, by performing studies with the previously generated apoC1-deficient mice. On a sucrose-rich, low fat/low cholesterol (LFC) diet, apoC1-deficient mice accumulate in their circulation VLDL particles, which contain relatively lower amounts of lipids when compared with VLDL isolated from control mice. Lipolysis assays in vitro on VLDL from apoC1-deficient and control mice showed no differences in apparent K(m) and Vmax values (0.27 +/- 0.06 versus 0.24 +/- 0.03 mmol of TG/litre and 0.40 +/- 0.03 versus 0.36 +/- 0.03 mmol of non-esterified fatty acid (NEFA)/min per litre respectively). To correct for potential differences in the size of the VLDL particles, the resulting K(m) values were also expressed relative to apoB concentration. Under these conditions apoC1-deficient VLDL displayed a lower, but not significant, K(m) value when compared with control VLDL (3.44 +/- 0.71 versus 4.44 +/- 0.52 mmol of TG2/g apoB per litre). VLDL turnover studies with autologous injections of [3H]TG-VLDL in vivo showed that the VLDL fractional catabolic rate (FCR) was decreased by up to 50% in the apoC1-deficient mice when compared with control mice (10.5 +/- 3.4 versus 21.0 +/- 1.2/h of pool TG). No significant differences between apoC1-deficient and control mice were observed in the hepatic VLDL production estimated by Triton WR139 injections (0.19 +/- 0.02 versus 0.21 +/- 0.05 mmol/h of TG per kg) and in the extra-hepatic lipolysis of VLDL-TG (4.99 +/- 1.62 versus 3.46 +/- 1.52/h of pool TG) in vivo. Furthermore, [125I]VLDL-apoB turnover experiments in vivo also showed a 50% decrease in the FCR of VLDL in apoC1-deficient mice when compared with control mice on the LFC diet (1.1 +/- 0.3 versus 2.1 +/- 0.1/h of pool apoB). When mice were fed a very high fat/high cholesterol (HFC) diet, the VLDL-apoB FCR was further decreased in apoC1-deficient mice (0.4 +/- 0.1 versus 1.4 +/- 0.4/h of pool apoB). We conclude that, in apoC1-deficient mice, the FCR of VLDL is reduced because of impaired uptake of VLDL remnants by hepatic receptors, whereas the production and lipolysis of VLDL-TG is not affected.

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Molecular characterization of quail apolipoprotein very-low-density lipoprotein II: disulphide-bond-mediated dimerization is not essential for inhibition of lipoprotein lipase.

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