A profile of the expression of a metabolic gene cluster in Arabidopsis

Plant cells often display a microtubule reorganization event when encountered with stress. This has been found to be integral for the reaction to stresses such as aluminum toxicity and cold stress. A cDNA microarray was previously conducted that identified MARNERAL SYNTHASE (MRN1), an oxidosqualene cyclase that produces the triterpene marneral, as the most highly upregulated gene when microtubule dynamics are disrupted in Arabidopsis. This work identifies two cytochrome P450s, CYP71A16 and CYP705A12, that are highly coregulated with MRN1 and are located within close proximity to the MRN1 loci. Using GC-FID and GC-MS, MRN1 and CYP71A16 are shown to function together in a single pathway in what is known as a metabolic gene cluster, while further testing shows that they are not in fact regulated by microtubule dynamics. The expression profile of these genes is explored since there is no known function for marneral or its related metabolites. Using a promoter-reporter and real time PCR analysis, it was found that the hormones ABA and methyl jasmonate induce expression of the three genes to different degrees depending on seedling age. Osmotic stressors, including mannitol and NaCl treatments, also induce the expression of these genes. MRN1, in particular, seems to show the highest level of induction suggesting that the pathway is transcriptionally regulated through MRN1. These conditions are shown to not affect the growth response in mutant plants unable to metabolize marneral or plants ectopically expressing different combinations of the three genes. These conditions are intriguing because most triterpenes derived from secondary metabolism are generally thought to play roles in defense, yet these data suggest that the pathway is induced under abiotic stress conditions. The marneral cluster may have evolved to be expressed under osmotic stress conditions in a sense to protect the plants water from pathogens or herbivores. It is also reasonable to speculate that these compounds may play roles in signalling or membrane modification. Further experiments are proposed that could test these hypotheses.Science, Faculty ofBotany, Department ofGraduat