Sorting and transepithelial transport of adsorbed protein tracers: effects of temperature.

Abstract

The epithelium of the guinea pig yolk sac is involved in the selective transport of macromolecules to the fetus. We studied the compartments involved in sorting and transepithelial transport of protein tracers and the effect of lowered temperature (18 degrees C) on these events. Explants of yolk sac were incubated with a mixture of cationized ferritin (CF) and horseradish peroxidase (HRP, Sigma type VI). At 4 degrees C, both tracers were bound to the cell surface and binding of an HRP-gold complex was shown to be inhibited by mannan. At 37 degrees C and 18 degrees C, both tracers were taken up into tubules and vesicles in the apical cytoplasm. Usually the tubules contained a mixture of tracers, but they often showed a polarized distribution with CF and HRP at opposite ends. The vesicles also contained mixtures of the tracers, but some contained only one. In addition, there were some irregularly shaped vacuoles composed of saccules that contained either a mixture, HRP alone, or CF alone. These results suggest that these adsorbed ligands are binding to unique microdomains of the endocytic complex. After 20 min at 37 degrees C coated vesicles 100 nm in diameter were located in the apical cytoplasm and coated vesicles of the same size were located at the lateral cell membrane. Usually they contained only HRP or CF, although occasional mixtures were seen. At 18 degrees C, HRP was transported across the cells in 100 nm vesicles. However, transport of CF was completely inhibited at the lower temperature.(ABSTRACT TRUNCATED AT 250 WORDS)