Cell damage by cytolysin. Spontaneous recovery and reversible inhibition by divalent cations.
Cytolysin-induced membrane damage (which requires low Ca2+) has been studied 1) in E by assay of hemolysis, 2) in Lettre cells by measurement of transmembrane potential, intracellular content of K+ and Na+, leakage of phosphoryl[3H]choline or 51Cr from [3H]choline-labeled or 51CrO4(2-)-labeled cells and leakage of lactate dehydrogenase, and 3) in phospholipid bilayers by measurement of electrical conductivity changes. In Lettre cells, damage is restricted and reversible: little lactate dehydrogenase leaks from cells that leak substantial amounts of Na+, K+, and phosphoryl[3H]choline; at low amounts of cytolysin, membrane potential and intracellular content of Na+ and K+ recover within minutes. In E and Lettre cells, membrane damage is inhibited by Zn2+, by high Ca2+, or by low pH. Inhibition is reversible: addition of EGTA to Zn2+-protected E or Lettre cells (incubated in the presence of cytolysin, low Ca2+ and Zn2+) initiates leakage; removal of Zn2+ (and cytolysin and Ca2+) by washing also initiates leakage; such leakage is again sensitive to Zn2+, high Ca2+, or H+. In phospholipid bilayers, channels induced by cytolysin (at low Ca2+) are partially closed by negative voltage; Ca2+, Zn2+, or H+ promote channel closure. Channels are re-opened (only partially in the case of Zn2+) by positive voltage. From all these results it is concluded that the action of cytolysin on membranes is similar to that of other pore-forming agents: damage does not necessarily lead to lysis of nucleated cells, and can be prevented by Ca2+, Zn2+, or H+.
Bashford
C L
CL
Department of Biochemistry, St. George's Hospital Medical School, London, UK.
Menestrina
G
G
Henkart
P A
PA
Pasternak
C A
CA
D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002286:Carcinoma, Ehrlich Tumor / Q000473:pathology; D002413:Cations, Divalent; D002470:Cell Survival / Q000187:drug effects*; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D008051:Lipid Bilayers / Q000378:metabolism; D008562:Membrane Glycoproteins; D008565:Membrane Proteins / Q000633:toxicity*; D051379:Mice; D054353:Perforin; D010743:Phospholipids / Q000378:metabolism; D052899:Pore Forming Cytotoxic Proteins; D011522:Protons; D051381:Rats; D015032:Zinc / Q000494:pharmacology
D002413:Cations, Divalent; D008051:Lipid Bilayers; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D010743:Phospholipids; D052899:Pore Forming Cytotoxic Proteins; D011522:Protons; D054353:Perforin; D015032:Zinc; D002118:Calcium