Uptake of high-density lipoprotein by Y-organs of the crab Cancer antennarius: III. Evidence for adsorptive endocytosis and the absence of lysosomal processing.
Ecdysteroid hormones in crustaceans are synthesized from cholesterol in the Y-organs. Circulating cholesterol is bound to high-density lipoprotein (HDL). Experiments were conducted to find the mode of cholesterol uptake by Y-organ cells. The working hypothesis is as follows: Cholesterol is taken up by endocytosis of the entire HDL-cholesterol complex. HDL was doubly labeled in the apolipoprotein and cholesterol components with 125I and 3H, respectively. The time courses of uptake of the two labels by Y-organ segments in vitro were parallel throughout 24 hr of incubation. Chloroquine, an inhibitor of lysosomal function, had no effect on the uptake and degradation of HDL. Cycloheximide, an inhibitor of protein synthesis, depressed HDL uptake. Transmission electron microscopy of Y-organ tissue showed features characteristic of adsorptive endocytosis, including coated pits in the plasma membranes and their invagination and vesiculation. Visualization of HDL with immunogold binding showed profiles consistent with endocytosis. Quantitation of the distribution of HDL-gold particles indicated that about 75% were associated with cellular formed elements, but not with lysosomes, and significantly more particles overall were present in cells from de-eyestalked crabs than those from intact crabs. These data demonstrate that (a) cholesterol-carrying HDL is taken into Y-organ cells by adsorptive endocytosis, (b) its uptake is dependent on de novo protein synthesis, (c) its intracellular processing is independent of lysosomal enzymes, and (d) its uptake is depressed by the molt-inhibiting hormone secreted by the eyestalks.