Endocytosis of cationized ferritin by soybean (glycine max L.) protoplasts
Protoplasts derived from soybean SB-I suspension cell cultures were incubated in buffered osmoticum containing cationized ferritin (CF) or native ferritin (NF) to demonstrate the process of endocytosis in plant protoplasts. Uptake of CF occurred rapidly; with as little as a 10 s incubation, CF was found in the cytoplasm in coated vesicles (CVs). After 30 s, CF was also found in small smooth (i.e. non-coated) vesicles. By 2 to 6 min, CF was occasionally detected in three additional organelles; the partially coated reticulum (PCR), dictyosomes and multivesicular bodies (MVBs). This is the first demonstration that the PCR is involved in the process of endocytosis in ptants. The PCR consists of- a network of interconnected tubular membraneswith a coat over part of its cytoplasmic surface. Serial sectioning established that CF was distributed throughout this organelle except for membrane dilations which occasionally contained small internal vesicles. The PCR was frequently, but not always, associated with dictyosomes. Only one example of a direct membrane connection between these organelles was observed. Longer CF incubations resulted in increased labelling of the PCR, dictyosomes, and MVBs. Incubations of 60 min or longer resulted in the labelling of membranous masses within vacuoles. The membranous masses resembled the contents of some CF-Iabelled MVBs and are likely derived from them. Endocytosis in soybean protoplasts was charge dependent since even at high concentrations, very little NF was internalized. Results from protoplasts post-fixed in zinc iodideosmium tetroxide (AIO) indicate that membrane connections exist between the endoplasmic reticulum and dictysdomes. The lack ZIO reaction product in some PCR is consistent with the proposed independence of this organelle from dictyosomes. In preliminary experiments, whole SB-1 cells were incubated in NF, CF and lanthanum chloride. Uptake of CF and lanthanum was observed but could not unambiguously be attributed to the occurrence of endocytosis. A scheme for the endocytotic pathway in soybean protoplasts is presented. The results from protoplasts are compared with those for receptor-mediated endocytosis in animal cells. The implications of this pathway are considered with respect to membrane recycling, molecular repair, degradation of macromolecules and receptor-mediated endocytosis in plant protoplasts and cells.
Doctor of Philosophy (Ph.D.)