Phosphate availability regulates ethylene biosynthesis gene expression and protein accumulation in white clover...

The expression and accumulation of members of the 1-aminocyclopropane-1-carboxylate (ACC) synthase (ACS) and ACC oxidase (ACO) gene families was examined in white clover roots grown in either P_i (phosphate) sufficient or P_i-deprived defined media. The accumulation of one ACO isoform, TR-ACO1, was positively influenced after only 1 h of exposure to low P_i, and this was maintained over a 7-day time-course. Up-regulation of TR-ACS1, TR-ACS2 and TR-ACS3 transcript abundance was also observed within 1 h of exposure to low P_i in different tissue regions of the roots, followed by a second increase in abundance of TR-ACS2 after 5-7 days of exposure. An increase in transcript abundance of TR-ACO1 and TR-ACO3, but not TR-ACO2, was observed after 1 h of exposure to low P_i, with a second increase in TR-ACO1 transcripts occurring after 2-5 days. These initial increases of the TR-ACS and TR-ACO transcript abundance occurred before the induction of Trifolium repens PHOSPHATE TRANSPORTER 1 (TR-PT1), and the addition of sodium phosphite did not up-regulate TR-ACS1 expression over 24 h. In situ hybridization revealed some overlap of TR-ACO mRNA accumulation, with TR-ACO1 and TR-ACO2 in the root tip regions, and TR-ACO1 and TR-ACO3 mRNA predominantly in the lateral root primordia. TR-ACO1p-driven GFP expression showed that activation of the TR-ACO1 promoter was initiated within 24 h of exposure to low P_i (as determined by GFP protein accumulation). These results suggest that the regulation of ethylene biosynthesis in white clover roots is biphasic in response to low P_i supply.