Uptake of the lipophilic cation dibenzyldimethylammonium into Saccharomyces cerevisiae. Interaction with the thiamine transport system

Barts, P.W.J.A.; Hoeberichts, J.A.; Klaassen, AndréB.M.; Borst-Pauwels, G.W.F.H.

doi:10.1016/0005-2736(80)90156-x

Cited by 53 publications

(16 citation statements)

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“…Although the validity of lipophilic cations to determine membrane potential quantitatively has been questionated in yeast, it is generally accepted that its accummulation in yeast, is a function of d y [8,[14][15][16][17][18][19]. Boxman et al [I41 have shown that the lipophilic cation Ph4P+ binds to several yeast constituents which invalidate this compound for quantitative determination of the membrane potential without correction for unspecific binding.…”

Section: Discussionmentioning

confidence: 99%

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Trehalase activation in yeasts is mediated by an internal acidification

et al. 1986

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It has been reported that the addition of glucose, uncouplers and nystatin to yeast cells grown in a sugarfree medium causes trehalase activation; it has been postulated that this activation might be mediated by the depolarization of the plasma membrane. In this article the values of membrane potential and pH gradient across the plasma membrane of Saccharomyces cerevisiae have been determined under the same conditions as those in which trehalase is activated. Membrane potential was evaluated from the distribution of triphenylmethylphosphonium, the pH gradient from the distribution of benzoic acid across the plasma membrane. When the effect of several agents on the two components of the electrochemical proton gradient across the plasma membrane of ethanol-grown yeast cells were studied, under trehalase activation conditions, the following observations were made. (a) The addition of glucose activated trehalase and caused internal acidification of the cells, but had practically no effect on the membrane potential. (b) The addition of 200 mM KCI depolarized the cell membrane but did not affect the internal pH, nor trehalase activity. (c) Although carbonyl cyanide mchlorophenylhydrazone depolarized the cells at external pH 6.0 and 7.0, it only activated trehalase at an external pH 6.0, leading to the acidification of the internal medium at this pH. (d) Nystatin caused an increase in the triphenylmethylphosphonium accumulation at external pH 6.0 and 7.0, but only activated trehalase at external pH 6.0, causing acidification of the cell interior at this pH. (e) Activation of trehalase was also observed when the internal acidification was caused by addition of a weak acid such as acetate. It is concluded that trehalase activation is mediated by an intracellular acidification and is independent of the membrane potential.It has been established by several authors that glucose, proton conductors and nystatin cause trehalose breakdown [l], gluconeogenesis inhibition [2, 31 and glycolysis stimulation [4] in Succharomyces cerevisiae. The regulatory effects of these agents is believed to be mediated by an increase in the levels of intracellular CAMP which leads to an activation of the CAMP-dependent protein kinase. This enzyme causes the subsequent activation of trehalase and phosphofructo-2-kinase and the partial inactivation of fructose-l,6-bisphosphatase among other enzymes [l -41. It has been postulated that the increase in internal CAMP is caused by the depolarization of the plasma membrane, since glucose, uncouplers and nystatin produce rapid depolarization in Neurospora crama [5, 61. Although these agents elicited an increase of the CAMP levels in several yeast strains [2 -51, proof of a direct relationship between changes in membrane potential ( A y ) and fluctuations in CAMP concentrations in S. cerevisiue is still lacking. Proton conductors and nystatin [7] are known to cause an influx of protons which could affect both parameters of the electrochemical proton gradient. Therefore, the increase in internal CAMP leve...

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Section: Discussionmentioning

confidence: 99%

“…Barts et al [16] have shown that this system is induced by growing the cells in glucose and that the uptake rate of dibenzyldimethylammonium' is inhibited by 90% in the presence of 1 FM thiamin. Similar competition is found between Ph3MeP+ and thiamin uptake.…”

Section: Discussionmentioning

confidence: 99%

Trehalase activation in yeasts is mediated by an internal acidification

et al. 1986

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“…Instead it utilizes a nonselective, conductive anion channel (Kramhoft et al, 1986). In this case, the steady-state transmembrahe distribution of SCN-should be governed exclusively by the chemical and electrical driving forces (Barts et al, 1980;Midgley & Thompson, 1985) and provide a measure of Vm. We have tested this expectation in a series of experiments designed to characterize the transport and effects of [t4C]SCN-in Ehrlich cells.…”

Section: Uptake and Accumulation Of [14c]scn By Ehrlich Cellsmentioning

confidence: 99%

Validation of the use of the lipophilic thiocyanate anion for the determination of membrane potential in Ehrlich ascites tumor cells

Smith

Robinson

1989

J. Membrain Biol.

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The utility of the lipophilic anion thiocyanate (SCN-) as a probe for the indirect estimation of the cell membrane potential (Vm) in Ehrlich ascites tumor cells has been evaluated by comparison to direct electrophysiological measurements. SCN accumulation is consistent with first-order uptake into a single, kinetically-identifiable cellular compartment, achieving steady-state distribution in 20-30 min at 22 degrees C. The steady state distribution ratio ([SCN-]e/[SCN-]e) in physiological saline is 0.44 +/- 0.02. Treatment of the cells with propranolol (0.13 mM), an activator of Ca2+ dependent K+ channels, reduces the steady-state distribution ratio to 0.19 +/- 0.02. Conversely, treatment with BaCl2 (10 mM), an antagonist of the pathway, increases the SCN- distribution ratio to 0.62 +/- 0.01. The equilibrium potentials (VSCN) calculated under these conditions are virtually identical to direct electrophysiological measurements of the Vm made under the same conditions. The effect of varying extracellular [K+] ([K+]e) in the presence of constant [Na+]e = 100 mM has also been tested. In control cells, elevation of [K+]e from 6 to 60 mM reduces VSCN from -20.6 +/- 1.0 to -13.2 +/- 1.2 mV. Again, microelectrode measurements give excellent quantitative agreement. Propranolol increases the sensitivity of the cells to varying [K+]e, so that a 10-fold elevation reduces VSCN by approximately 31 mV. BaCl2 greatly reduces this response: a 10-fold elevation in [K+]e yielding only a 4-mV reduction in VSCN. It is concluded that the membrane potential of Ehrlich cells can be estimated accurately from SCN- distribution measurements.

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“…The reason for the disagreement between the values calculated using the data from Cs+-valinomycin experiments and those obtained using methyltriphenylphosphonium ion is currently unknown but there have been a number of reports indicating limitations to the use of this probe in other systems (Barts et al, 1980;Lolkema et al, 1982). Values obtained using the valinomycin system are usually considered to reflect the true value more closely as significant intracellular binding of inorganic cation is considered not to occur, in contrast to the behaviour of the lipophilic cations (Rink, 1982).…”

Section: Discussionmentioning

confidence: 91%