Vacuolar ethylene formation does not depend on membrane potential

Enzymatic synthesis of ethylene in the vacuole is assumed to require membrane integrity. The possibility that this reflects dependence on the vacuolar membrane potential was investigated. Vacuoles were released from protoplasts isolated from leaves of Vicia faba L. cv. Cyprus. The dependence of the ethylene-forming activity on tonoplast integrity was re-examined by immobilization of the vacuoles in a cross-linked polymeric matrix and subsequent permeabilization of the tonoplast with toluene, a pore-forming reagent. The relationship between the vacuolar ethylene formation and the membrane potential of free vacuoles was investigated by following the uptake of thiocyanate using permeabilized, depolarized and hyperpolarized vacuoles. Toluene and the proton conductor carbonyl cyanide m -chlorophenylhydrazone (CCCP) caused loss of ethylene-forming activity and depolarized the vacuolar membrane potential. However, depolarization of the membrane potential with choline chloride and hyperpolarization by ATP did not affect ethylene biosynthesis. These conflicting results lead to the conclusion that vacuolar ethylene biosynthesis is not dependent on the vacuolar membrane potential. The possibility that the inhibition of ethylene biosynthesis by toluene and CCCP may result from direct hydrophobic interactions between these compounds and hydrophobic components of the ethylene-forming enzyme is discussed.