Journal of Lipid Research, Vol 34, 1093-1106, Copyright © 1993 by Lipid Research, Inc.

ARTICLES

High density lipoprotein3 binding sites are related to DNA biosynthesis in the adenocarcinoma cell line A549

G Favre, KA Tazi, F Le Gaillard, F Bennis, H Hachem and G Soula
Laboratoire Ciblage en Therapeutique, Faculte des Sciences Pharmaceutiques et Centre Claudius Regaud, Toulouse, France.

The effect of high density (apoE-depleted HDL3) on cell growth of a human tumor cell line (A549) was studied and related to its binding on the plasma membrane. HDL3 were shown to stimulate the incorporation of [3H]thymidine into DNA and cell proliferation; these effects were dose- dependent. As HDL3, apoA-I and apoA-I-liposomes complexes (but not apoA- II) were able to stimulate DNA synthesis in serum-free conditions. This effect was maximum for 15-30 micrograms HDL3 protein/ml concentration. Binding of HDL3 on whole cells occurred by two mechanisms: the first was specific for HDL3; the second, of lower affinity, was phospholipid- dependent and was inhibited by low density lipoprotein or by phospholipid particles. Internalization and degradation of bound HDL3 were not observed. The specific sites (27.9 +/- 2.2 ng HDL3 protein/ng cell protein) accounted for only 2.5% of total (specific+phospholipid) binding sites and they bound HDL3 with a dissociation constant (KD) of 2.47 +/- 0.46 microgram HDL3 protein/ml (2.6 +/- 0.5 x 10(-8) M). The apparent KD value of total binding sites (specific+phospholipid) was eightfold higher (20.4 +/- 6.1 micrograms HDL3 protein/ml). Analysis of the membrane specific binding sites by ligand blotting with 125I- labeled HDL3 showed a single protein with an apparent molecular mass of 110 kDa. When HDL3 binding on phospholipid sites was inhibited by rigid phospholipid particles, the stimulation of [3H]thymidine incorporation related to HDL3 concentration did not show a maximum peak as previously observed but reached a plateau at a concentration as low as 5 micrograms HDL3 protein/ml. This low concentration also nearly saturated the specific binding sites with HDL3. When binding on specific protein sites was suppressed by tetranitromethane, DNA synthesis was not stimulated but, in contrast, inhibited. The stimulating effect of HDL3 on DNA biosynthesis is therefore likely dependent on HDL3 occupying specific binding sites.
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