Journal of Lipid Research, Vol 33, 233-240, Copyright © 1992 by Lipid Research, Inc.

ARTICLES

Identification of a neutral lipid core in a transiently expressed and secreted lipoprotein containing an apoB-48-like apolipoprotein

DJ Spring, SM Lee, DL Puppione, M Phillips, J Elovson and VN Schumaker
Department of Chemistry and Biochemistry, University of California, Los Angeles.

The presence of core lipids in lipoproteins expressed and secreted by transfected HepG2 cells was demonstrated by measuring the densities of these lipoproteins before and after treatment with a bacterial lipase specific for neutral lipids. HepG2 cells were reproducibly transfected with pRSV/B48, containing a truncated human apolipoprotein B-100 (apoB- 100) cDNA (nucleotides 1 to 6860, where nucleotide 129 is the start of translation). Northern blots of cellular message probed with apoB-48 showed abundant transcription of an apoB-48-sized message as well as endogenous apoB-100 message. When grown in the presence of [35S]methionine, pRSV/B48-transfected cells secreted lipoproteins containing an apoB-48-like apolipoprotein. This lipoprotein banded at a density of 1.11 g/ml in isopycnic NaBr gradients. Electron microscopy of the apoB-48-containing lipoproteins demonstrated spherical particles with an average diameter of 124A. A sedimentation rate of 8.4S was measured by sucrose gradient sedimentation. When the apoB-48-containing particles were treated with a bacterial lipase (from Chromobacterium viscosum), shown to hydrolyze triglycerides and cholesteryl esters but not phospholipids, their density increased to 1.18 g/ml, consistent with removal of core lipids. When the secreted lipoprotein was modeled as a spherical particle containing a single molecule of apoB-48, a triglyceride-filled core, and a surface monolayer of phospholipid and protein, the hydrodynamic properties were consistent with the observed sedimentation coefficient, buoyant densities before and after lipase treatment, and the diameter as seen with the electron microscope. These data indicate that transfected HepG2 cells assembled and secreted lipoproteins possessing the same physical structure as naturally occurring lipoproteins.
CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				R. B. Weinberg, V. R. Cook, J. A. DeLozier, and G. S. Shelness Dynamic interfacial properties of human apolipoproteins A-IV and B-17 at the air/water and oil/water interface J. Lipid Res., September 1, 2000; 41(9): 1419 - 1427. [Abstract] [Full Text]

				Q. Xiao, J. Elovson, and V. N. Schumaker Rat McA-RH7777 cells efficiently assemble rat apolipoprotein B-48 or larger fragments into VLDL but not human apolipoprotein B of any size J. Lipid Res., January 1, 2000; 41(1): 116 - 125. [Abstract] [Full Text]

				A.-M. Brown, D. Wiggins, and G. F. Gibbons Glucose Phosphorylation Is Essential for the Turnover of Neutral Lipid and the Second Stage Assembly of Triacylglycerol-Rich ApoB-Containing Lipoproteins in Primary Hepatocyte Cultures Arterioscler. Thromb. Vasc. Biol., February 1, 1999; 19(2): 321 - 329. [Abstract] [Full Text] [PDF]

				E. Nicodeme, F. Benoist, R. McLeod, Z. Yao, J. Scott, C. C. Shoulders, and T. Grand-Perret Identification of Domains in Apolipoprotein B100 That Confer a High Requirement for the Microsomal Triglyceride Transfer Protein J. Biol. Chem., January 22, 1999; 274(4): 1986 - 1993. [Abstract] [Full Text] [PDF]

				P. J. Coussons, C. S. Bourgeois, D. Wiggins, and G. F. Gibbons Selective Recruitment of ApoB-48 for the Assembly of VLDL in Rat Triacylglycerol-Enriched Hepatocytes Arterioscler. Thromb. Vasc. Biol., July 1, 1996; 16(7): 889 - 897. [Abstract] [Full Text]

				T. L. Innerarity, J. Borén, S. Yamanaka, and S.-O. Olofsson Biosynthesis of Apolipoprotein B48-containing Lipoproteins J. Biol. Chem., February 2, 1996; 271(5): 2353 - 2356. [Full Text] [PDF]