Differences in endocytosis and intracellular sorting of ricin and viscumin in 3T3 cells.
Ricin and viscumin are heterodimeric protein toxins. Their A-chain is enzymatically active and removes an adenine residue from the 28S rRNA, the B-chain has lectin activity and binds to terminal galactose residues of cell surface receptors. The toxins reveal a high degree of identity in their amino acid sequences. Nevertheless, uptake into 3T3 cells occurs via different receptors and endocytotic pathways. This has been revealed by enzyme linked based analysis of ricin competition with viscumin, and by fluorochrome-labeled toxins (viscumin-FITC, ricin-Alexa 568), which were added simultaneously or separately to living cells. Then the uptake was followed by confocal laser scanning microscopy. Ricin immediately is delivered to the tubular and vesicular structures of endosomes in the perinuclear area while viscumin becomes endocytosed into small vesicles preferentially in the cell periphery. After about 60 min both these toxins may be found in tubo-vesicular structures of endosomes where the sorting process can directly be observed. The fact that this sorting takes place is a strong argument for the assumption that the toxins are bound to membrane proteins, either to their original receptors or to other proteins inside the endosomal compartment exhibiting terminal galactose residues. The toxins are biologically fully active as has been proven by binding and by toxicity experiments, thus the differences in targeting do not arise from labeling.
Moisenovic
Mihail
M
Department of Cell Physiology and Immunology, Biological Faculty, MV Lomonosov Moscow State University, Russia.
Tonevitsky
Alexandr
A
Agapov
Igor
I
Niwa
Hideaki
H
Schewe
Heinz
H
Bereiter-Hahn
Jürgen
J
D016475:3T3 Cells; D000818:Animals; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D004705:Endocytosis / Q000187:drug effects* / Q000502:physiology; D011992:Endosomes / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D005057:Eukaryotic Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D006056:Golgi Apparatus / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D007425:Intracellular Membranes / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D051379:Mice; D028321:Plant Preparations / Q000378:metabolism / Q000494:pharmacology*; D010940:Plant Proteins; D011500:Protein Synthesis Inhibitors / Q000378:metabolism / Q000494:pharmacology*; D011990:Receptors, Transferrin / Q000187:drug effects / Q000378:metabolism; D054790:Ribosome Inactivating Proteins, Type 2; D012276:Ricin / Q000378:metabolism / Q000494:pharmacology*; D013997:Time Factors; D014118:Toxins, Biological / Q000378:metabolism / Q000494:pharmacology*; D022161:Transport Vesicles / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure
D028321:Plant Preparations; D010940:Plant Proteins; D011500:Protein Synthesis Inhibitors; D011990:Receptors, Transferrin; D054790:Ribosome Inactivating Proteins, Type 2; D014118:Toxins, Biological; C009674:ribosome inactivating protein, Viscum; D012276:Ricin