Enlargeosome traffic: exocytosis triggered by various signals is followed by endocytosis, membrane shedding or both.
Enlargeosomes are cytoplasmic organelles discharged by regulated exocytosis, identified by immunofluorescence of their membrane marker, desmoyokin/Ahnak, but never revealed at the ultrastructural level. Among the numerous enlargeosome-positive cells, the richest and most extensively characterized are those of a PC12 clone, PC12-27, defective of classical neurosecretion. By using ultrastructural immunoperoxidase labeling of formaldehyde-fixed, Triton-X-100-permeabilized PC12-27 cells, we have now identified the enlargeosomes as small vesicles scattered in the proximity of, but never docked to, the plasma membrane. Upon stimulation, these vesicles undergo exocytosis [rapid after the Ca(2+) ionophore, ionomycin, much slower after either the phorbol ester, phorbol myristate acetate (PMA), or ATP, working through a P2Y receptor], with appearance in the plasma membrane of typical desmoyokin/Ahnak (d/A)-positive, Omega-shaped and open profiles evolving into flat patches. Postexocytic removal of the exocytized d/A-positive membrane occurs by two processes: generation of endocytic vesicles, predominant after ionomycin and ATP 100-500 microM; and shedding of membrane-bound cytoplasmic bodies, predominant after PMA and 1 mM ATP, containing little or no trace of endoplasmic reticulum, Golgi, endo/lysosomes and also of a plasma membrane marker. Depending on the stimulation, therefore, the cell-surface expansion by enlargeosome exocytosis is not always recycled but can induce release of specific membranes, possibly important in the pericellular environment.
Cocucci
Emanuele
E
Center of Excellence in Cell Development, Vita-Salute San Raffaele University, DIBIT, via Olgettina 58, 20132 Milan, Italy.
Racchetti
Gabriella
G
Podini
Paola
P
Meldolesi
Jacopo
J
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D002999:Clone Cells; D003902:Detergents / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004705:Endocytosis / Q000187:drug effects / Q000502:physiology*; D011992:Endosomes / Q000187:drug effects / Q000648:ultrastructure; D005089:Exocytosis / Q000187:drug effects / Q000502:physiology*; D007425:Intracellular Membranes / Q000187:drug effects / Q000648:ultrastructure; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D016716:PC12 Cells; D051381:Rats; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D003902:Detergents; D007476:Ionophores; D015759:Ionomycin; D000255:Adenosine Triphosphate; D013755:Tetradecanoylphorbol Acetate