Ethylene is a major stimulating factor for natural rubber production in Hevea brasiliensis and is often applied in the form of Ethephon, an ethylene releaser. In a positive feedback type mechanism, the application of ethylene leads to the enhanced expression of genes involved in ethylene biosynthesis such as ACC oxidases, which are also responsive to wounding. We studied the regulation of ethylene biosynthesis in rubber tree through the characterization of the ACC oxidase multigene family and the responses of individual genes to ethylene and wounding. Three full-length cDNAs, HbACO1 (EMBL accession AM743170), HbACO2 (EMBL accession AM743171), and HbACO3 (EMBL accession AM743172) were isolated. All of the HbACO genes were expressed at all stages of development studied, from in vitro callus to the exploited plant, but with different expression profiles (Kuswanhadi et al. 2005). HbACO1 was expressed at a higher level than the other genes, and was down-regulated by ethylene and wounding in both leaf and bark tissues. In contrast, HbACO2 and HbACO3 were transiently induced in response to ethylene and wounding. Treatment with 1-MCP, an ethylene action inhibitor, abolished the ethylene induction of HbACO2 expression, showing that this gene was under positive feedback regulation. The expression of HbACO3 was also induced by ethylene treatment and wounding, again with an antagonistic effect of 1-MCP. In conclusion, HbACO1 appears to be responsible for basal levels of ethylene production while HbACO2 and HbACO3 are up-regulated in response to external factors.