calcimycin and 2--4--dimethylbenzamil

Streptococcus pneumoniae requires 0.15 mM-Ca2+ in the medium for optimal growth. Increasing the Ca2+ concentration to 1 mM triggers either a differentiative state, competence for genetic transformation during exponential growth, or partial lysis as soon as the cultures enter stationary phase. Genetic and physiological data both suggest that these responses are under the control of activator(s), excreted in the presence of high Ca2+ concentrations. 45Ca2+ transport is also stimulated by the activator(s). The amiloride derivative 2',4'-dimethylbenzamil (DMB) inhibits 45Ca2+ transport and prevents lysis and competence development. This provides evidence in favour of the involvement of Ca2+ transport in competence and culture lysis. On the other hand, addition of DNA to a competent culture prevents lysis of wild-type bacteria while a mutant, defective for DNA uptake, is not protected from lysis by exogenous DNA. An hypothesis is proposed for competence induction as a global metabolic response to Ca2+, under the control of competence factor.

Topics: Amiloride; Bacteriolysis; Biological Transport; Calcimycin; Calcium; Culture Media; DNA; Mutation; Potassium; Sodium; Streptococcus pneumoniae; Transformation, Genetic

In NK cell-mediated cytolysis, the degranulation of the NK cell, and the binding and polymerization of its effector molecule cytolysin/perforin, are Ca2+ dependent. To determine if increases in the target cell free intracellular Ca2+ concentration ([Ca2+]i) are also important for cytotoxicity, we examined the effects of Ca2+ substitutes, promotors of Ca2+ influx; and inhibitors of Ca2+ efflux on the cytolysis mediated by purified cytolysin. Ca2+ promoted cytolysin-mediated cytolysis but could also inactivate cytolysin on preincubation in the absence of target cells. Ba2+ and Sr2+ could not alone promote cytolysin-mediated cytolysis but did enhance Ca2(+)-driven cytolysis. Preincubation with Ba2+ and Sr2+ did not inactivate cytolysin. One interpretation which these results suggested was that neither Ba2+ nor Sr2+ were involved in cytolysin polymerization and pore formation, but could act intracellularly as Ca2+ analogs once pore formation had occurred. The synergistic interaction of cytolysin and the Ca2+ ionophore A23187 further supported a role for increased [Ca2+]i in cytolysis. Inhibitors of Na+/Ca2+ exchange, namely low Na+ medium, ouabain, and the amiloride analogs 2',4'-dimethylbenzamil, 5-(N-4-chlorobenzyl)-2',4'dimethylbenzamil, and alpha-phenylbenzamil all markedly enhanced cytolysin-mediated cytolysis. These results support the hypothesis that the Ca2+ dependency of NK cell- and cytolysin-mediated cytolysis is related to increases in target cell [Ca2+]i. Furthermore, they indicate that Na+/Ca2+ exchange is an important counterlytic mechanism.

Topics: Amiloride; Animals; Barium; Biological Transport; Calcimycin; Calcium; Carrier Proteins; Cell Line; Cell Survival; Cytotoxicity, Immunologic; Cytotoxins; Killer Cells, Natural; Membrane Glycoproteins; Membrane Proteins; Mice; Ouabain; Perforin; Pore Forming Cytotoxic Proteins; Sodium; Sodium-Calcium Exchanger; Sodium-Potassium-Exchanging ATPase; Strontium; Verapamil