Expression of carboxyl-terminally truncated forms of human apolipoprotein B in rat hepatoma cells. Evidence that the length of apolipoprotein B has a major effect on the buoyant density of the secreted lipoproteins.
We examined the relationship between the size of human apolipoprotein (apo) B and the formation and secretion of apoB-containing lipoprotein particles. Stable transformants of the rat hepatoma cell line McA-RH7777 harboring a variety of human apoB cDNA constructs were established, and these produced carboxyl-terminally truncated apoB proteins (apoB18, -B23, -B28, -B31, -B48, and -B53). Immunoblotting of apoB proteins secreted into the culture medium and fractionated by equilibrium density ultracentrifugation revealed that each of the truncated apoB species was secreted from the cells. The peak densities of the apoB-containing particles decreased as the length of the apoB proteins increased. Apolipoproteins B18 and B23 appeared at the bottom of the salt gradient (d = 1.23 g/ml), whereas particles containing apoB28, -B31, -B37, -B48, and -B53 exhibited progressive decreases in density. The density distribution of secreted apolipoproteins was not affected by the expression or secretion of these recombinant apoB species. As determined by nondenaturing gel electrophoresis, apoB28, -B31, -B37, -B48, and -B53 formed their own discrete particles, and there was a direct correlation between the size of the particles and the length of the apoB species. The efficiency and rate of secretion of these truncated forms of apoB were studied by measuring the decrease of immunoprecipitated 35S-labeled apoB proteins in the cells and their accumulation in the medium. Proteins corresponding to apoB28 or larger were rapidly and efficiently secreted, whereas apoB18 and apoB23 were secreted much more slowly. These data imply that the size of these truncated apoB forms governs the lipid content of the apoB-containing lipoproteins formed as well as the kinetics of secretion.
Yao
Z M
ZM
Gladstone Foundation Laboratories for Cardiovascular Disease, University of California, San Francisco 94140-0608.
Blackhart
B D
BD
Linton
M F
MF
Taylor
S M
SM
Young
S G
SG
McCarthy
B J
BJ
D000818:Animals; D001055:Apolipoproteins B / Q000096:biosynthesis / Q000235:genetics*; D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007700:Kinetics; D008114:Liver Neoplasms, Experimental / Q000378:metabolism*; D010957:Plasmids; D051381:Rats; D014162:Transfection; D014407:Tumor Cells, Cultured
D001055:Apolipoproteins B