Characterization and Expression Pattern of 1-Aminocyclopropane-1-Carboxylate (ACC) Synthases cDNA from Geranium (Pelargonium x hortorum cv Sincerity)

Abstract

The enzyme 1-aminocyclopropane-1-caryboxylate (ACC) synthase catalyzes the rate limiting step in the regulation of ethylene biosynthesis and is, primarily, regulated at the transcription level. To understand the role of ACC synthases in stress-induced ethylene bursts in geranium, degenerated oligonucleotide primers were used to amplify a DNA probe from cDNA; and used to screen a cDNA library prepared from an mRNA preparation from mature flowers. Six putative full-length ACC synthase clones were isolated; these clones were divided into two groups on the basis of deduced amino acid sequences and features contained in their mRNA. Clones (e.g. PHS-ACS41, 44, and 45) in group I contain a stop codon (TAG) just before the initiation codon (ATG). Clone PHS-ACS44 from this group contains an “extra” 780 nucleotide sequence at the untranslated 3′-end region (UTR) after the first polyadenylation signal and a short poly(A) tail. The “extra” UTR contains another polyadenylation signal and a poly(A) tail, otherwise the coding sequence is identical to PHS-ACS41. Clone PHS-ACS49 in group II contains stop codon (TAG) twenty one bases upstream from the initiation codon. All these genes contain eleven conserved amino acid residues, the substrate and pyridoxal 5′-phosphate binding site that are characteristic of all ACC synthases. Using RTPCR and gene specific primers, expression studies show that these genes are expressed differentially. PHS-ACS41 and PHSACS44 not only show differences in expression in vivo, but also exhibit differential translational efficiencies in vitro suggesting that the “extra” 780 UTR in vivo may serve as a regulatory element. Our finding of multiple ACC synthase genes and their expression in geranium is consistent with similar observations in other plant species in which ACC synthases are encoded by a multigene family that are differentially regulated by internal and external factors.