Characterization of 1-Aminocyclopropane-1-Carboxylate Deaminase Containing Methylobacterium spp. Isolated from Rhizosphere Soils of Field-Grown Rice and Regulation of Ethylene Levels in Canola.
Munusamy Madhaiyan1, Selvaraj Poonguzhali1, Jeounghyun Ryu1, Woojong Yim1, Myoungsu Park2, and Tongmin Sa1. (1) Chungbuk National University, 12, Gaesin-dong, Heungduck-ku, Cheongju, South Korea, (2) Semyung University, San 21-1, Shinwol-dong, Jecheon, South Korea
The plant hormone ethylene is of great importance in plant growth and development. An increased concentration of endogenous ethylene in plants or an enhanced sensitivity of certain plant species, mainly dicots, to ethylene can result in inhibition of seed germination and root growth. One of the mechanisms that a number of plant growth promoting bacteria use to facilitate plant growth and development is the lowering of a plant's ethylene concentration through the action of the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase, that is quite effective in lowering the concentration of ACC and hence ethylene in plant tissues. The presence of ACC deaminase has been studied in various plant growth promoting bacteria (PGPB) like Enterobacter cloacae, Rhizobium, and Pseudomonas, Variovorax, Alcaligenes, Bacillus etc. The ACC deaminase containing bacteria may be found on leaves and flowers as well as on seeds and roots, and the model based on the interaction of bacteria with seeds and roots is likely applicable to the entire plant. Bacteria of the genus Methylobacterium (PPFMs – Pink- pigmented facultative methylotrophic bacteria) are strict aerobic, Gram-negative rods, able to grow on C1 compounds. Several aspects of plant growth promotion by Methylobacterium have also been investigated, such as the production of urease enzyme, stimulation of seed germination, promotion of root growth and morphology and induced systemic resistance. In this study, we surveyed Methylobacterium strains for ACC deaminase activity and the various steps involved in lowering the ethylene levels as proposed in the model for plant growth promotion by ACC deaminase containing PGPB. Here, we report the presence of ACC deaminase in Methylobacterium fujisawaense and it's lowering of ethylene levels and promotion of root elongation in canola seedlings under gnotobiotic conditions. Three of the five Methylobacterium strains used in our study were found to possess ACC deaminase activity and they were selected for growth pouch assays and for testing plant growth promoting traits in canola. To test a part of the previous model proposed for ACC deaminase producing bacteria with Methylobacterium, ACC levels and various enzyme activities were monitored in canola. Lower amounts of ACC were present in the tissues of seeds treated with M. fujisawaense strains than in control seeds treated with MgSO4. Though the increased activities of ACC synthase in the tissue extracts of the treated seedlings might be due to bacterial indole-3-acetic acid (IAA), the amount of ACC was reduced due to bacterial ACC deaminase activity. The activities of ACC oxidase, the enzyme catalyzing conversion of ACC to ethylene remained lower in M. fujisawaense treated seedlings. This consequently lowered the ethylene in plants and prevented ethylene inhibition of root elongation. The results obtained here suggests that plant growth promotion by the ACC utilizing Methylobacterium is due to the reduction of plant ethylene through the utilization of ammonia cleaved from ACC by its ACC deaminase and this is the first report on the existence and prevalence of ACC deaminase in Methylobacterium spp. providing evidence for promotion of root elongation. From our results, we proposed a model for how Methylobacterium promote plant growth. Inoculation of canola with Methylobacterium strains increases the IAA and cytokinin concentration of the plants resulting in increased ACC synthase (ACS) activity. If the PGPR strain contains no ACC deaminase, ACC levels in the seedlings is unaltered and ethylene is still produced which inhibits root elongation. If the PGPR strain contains ACC deaminase, ACC levels in the plant drop reducing ethylene levels and releasing root growth from the inhibitory effects of this hormone. Also the ubiquitous nature and the ability to effectively utilize methanol, the metabolic waste product of plants provides additional benefits the use of Methylobacterium as a plant growth promoting bacteria.