Chilling-Induced Inactivation and Its Recovery of Tonoplast H⁺-ATPase in Mung Bean Cell Suspension Cultures

Abstract

The processes involved in adaptation to cold temperature were examined by growing suspension cultured cells of mung bean (Vigna radiata [L.] Wilczek) at 2 degrees C for various periods of time and assaying the activities of various membrane-bound enzymes in vitro. The tonoplast H(+)-ATPase activity and the ATP-proton transport extracted from cells incubated at 2 degrees C declined rapidly and reached a minimum level after 10 hours. The inactivation was reversible within 24 hours of chilling. The recovery of the cold-inactivated H(+)-ATPase was found to proceed in two steps, a faster recovery of ATP hydrolysis activity and a slower recovery of the proton transport. The recovery was markedly inhibited by the presence of azide, but not affected by 0.578 millimolar cycloheximide. This suggested the involvement of an energy process that had no requirement for de novo synthesis of protein. The cold-induced inactivation of the H(+)-ATPase may be due to a structural alteration of the enzyme. The slower recovery of proton transport relative to ATP hydrolysis during warming suggests that the protogenic domains in the enzyme may be affected differently by chilling.