The efficiency and kinetics of secretion of apolipoprotein A-I in hepatic and non-hepatic cells

Abstract

Apo A-I, the major protein component of high density lipoprotein (HDL), is synthesized by hepatic and intestinal cells and assembled with lipids to produce, in as yet incompletely understood ways, a mature HDL particle. For many secreted proteins only a portion of newly synthesized polypeptides are secreted, with the remainder being degraded at intracellular sites. For example apolipoprotein B secretion is controlled by the extent of intracellular degradation of the protein. Here we have systematically examined whether there is significant intracellular degradation of nascent apo A-I. We find that in two hepatic cell types, primary cultures of hepatocytes from cynomolgus monkey and HepG2 hepatocarcinoma cells, essentially all apo A-I that is synthesized is eventually secreted. A non-hepatic cell line, Chinese hamster ovary cells transfected with the apo A-I gene, secreted somewhat less (65%) of the apo A-1 synthesized. In a careful kinetic analysis, the rate of apo A-I secretion was found to be identical between the three cell types. This indicates that the mechanisms governing secretion are conserved among the different cell types. Further, the rate of secretion was the same for apo A-I in a lipid-poor form and in a form found associated in the medium with sufficient lipid to promote flotation in density gradients. The kinetic analysis indicates that there are two rate limiting steps to apo A-I secretion from the cell. It has previously been suggested that, for most proteins, exit from the endoplasmic reticulum is the rate limiting step in the secretory pathway. However, the first of the measured rate constants does not correspond to previously measured rates of exit from the endoplasmic reticulum, indicating that exit from the endoplasmic reticulum is probably not rate limiting in apo A-I secretion. Together, these data suggest that in hepatic cells, contrary to what is observed for apo B secretion, intracellular degradation and transport are not limiting factors in apo A-I secretion.