1-AMINOCYCLOPROPANE-1-CARBOXYLIC ACID-INDUCED ETHYLENE STIMULATES CALLUS FORMATION BY CELL ENLARGEMENT IN THE CAMBIAL REGION OF INTERNODAL EXPLANTS OF CITRUS

The effect of 1-aminocyclopropane-1-carboxylic acid (ACC) treatments o n callus formation from internodal explants of Troyer citrange (Citrus sinensis (L.) Osb. x Poncirus trifoliata) was investigated and the ce llular types responsible of callus formation were identified. The in v itro culture of explants in media supplemented with 0.05, 0.5 and 2 mM ACC resulted in rapid growth of whitish soft friable calli, whereas n o calli were observed in the absence of ACC. The calli were evident 3- 5 days after culture and callus growth reached a stationary phase 11 d ays after culture. A rapid and transitory increase in the rate of ethy lene production occurred at the highest ACC concentrations used. Callu s formation correlated with amounts of ethylene released by the explan ts. The lowest ACC concentration (0.05 mM) stimulated callus growth in only 10-20% of the explants, with callus weighing 100 mg, and the exp lants produced low ethylene levels at day 3 of culture (33 nl g(-1) FW h(-1)). The highest ACC concentrations (0.5 and 2 mM) stimulated call us growth in 60-75% of the explants, with callus weights of 155 and 21 0 mg, respectively. The explants produced high levels of ethylene at d ay 3 of culture (330-430 nl g(-1) FW h(-1)). The inhibition of ethylen e biosynthesis or action with cobalt chloride and silver thiosulfate, respectively, reduced callus formation in the ACC-treated explants in a concentration-dependent manner. Taken together, these observations i ndicate that ethylene synthesized from ACC is responsible for callus f ormation. Light-microscopy observations of the explants cultured in AC C showed that ethylene promoted enlargement of the procambial and pare nchymatous cells located between the primary phloem fibers and the met axylem. The examination of the micrographs also suggested that ethylen e-induced cell growth is mostly due to radial expansion. Thus, ACC-ind uced ethylene is responsible for the enlargement of the procambial and parenchymatous cells from Citrus internodal explants that, in turn, r esults in the stimulation of callus formation.