Autoregulation of the gene for cystathionine γ-synthase in Arabidopsis: post-transcriptional regulation induced by S-adenosylmethionine

Abstract

Cystathionine gamma-synthase (CGS) catalyses the first committed step of methionine biosynthesis in higher plants. CGS is encoded by the CGS1 gene in Arabidopsis. Stability of CGS1 mRNA is down-regulated in response to methionine application and the exon 1-coding region of CGS1 itself is necessary and sufficient for this regulation. mto1 (for methionine overaccumulation) mutants of Arabidopsis, which carry single-amino-acid sequence alterations within CGS1 exon 1, are deficient in this regulation and overaccumulate methionine. Since CGS1 exon 1 acts in cis during this regulation, we have proposed a model that the regulation occurs during translation of CGS1 mRNA when the nascent polypeptide of CGS and its mRNA are in close proximity. In fact, application of the translation inhibitor cycloheximide abolished this regulation in vivo. This model predicts that the regulation can be reproduced in an in vitro translation system. Studies using the in vitro translation system of wheatgerm extract have indicated that S-adenosylmethionine, a direct metabolite of methionine, is the effector of this regulation. A 5'-truncated RNA species, which is a probable degradation intermediate of CGS1 mRNA in vivo, was also detected in vitro, suggesting that the wheatgerm in vitro translation system reflects the in vivo regulation.

Aspartate-Derived Amino Acid Biosynthesis inArabidopsis thaliana

“…Additional mutational analysis of the At3g01120 N-terminus defined a regulatory domain of 11 to 13 amino acids that is highly conserved among plants and controls cystathionine γ-synthase mRNA stability (Ominato et al, 2002). Experiments with an in vitro translation system identified S-adenosylmethionine, rather than methionine, as the effector that binds to the N-terminus of cystathionine γ-synthase to reduce mRNA stability Onouchi et al, 2004). Binding of S-adenosylmethionine was also shown to cause translational arrest (Lambein et al, 2003;Onouchi et al, 2005), which precedes mRNA decay and is associated with the formation of a series of 5'-truncated mRNA products that may reflect the spacing of ribosomes on the mRNA (Haraguchi et al, 2008).…”

Nascent Peptide-dependent Translation Arrest Leads to Not4p-mediated Protein Degradation by the Proteasome

“…Rmp1p is a cytoplasmic component of the RNase MRP (31), an endonuclease that contributes to cell cycle-regulated degradation of daughter cell-specific mRNA (32). Because translation arrest leads to endonucleolytic cleavage of mRNA in prokaryotes and eukaryotes (8,9,(33)(34)(35)(36), translation arrest induced by consecutive basic amino acid sequences may lead to an endonucleolytic cleavage of mRNA, and RNase MRP may be responsible for this cleavage activity. It is likely that the cleavage products are degraded from the 3Ј end by an exosome or from the 5Ј end by Xrn1p and would only be detected in mutant cells.…”

“…The interaction between the SecM nascent peptide and the wall of the ribosomal exit tunnel functions as a discriminating gate during translation (6,7). Several eukaryotic examples of translation arrest also have been described (8,9). Moreover, the synthesis of polylysine caused by translation of a poly(A) sequence leads to translation arrest (3,4).…”

Improving the levels of essential amino acids and sulfur metabolites in plants

“…This chloroplast-localised enzyme uses O-phosphohomoserine and cysteine to form cystathionine, the first intermediate metabolite of the Met biosynthesis pathway (Figure 1). CGS is probably not regulated through feedback inhibition of enzyme activity (Ravanel et al, 1998a,b); instead, the amount of CGS enzyme is regulated by free Met through its downstream product, SAM, by a post-transcriptional mechanism (Chiba et al, 1999Onouchi et al, 2004). This regulation is associated with a specific domain in the CGS mRNA encoding the ca.…”

“…The regulatory significance of this domain was first identified through genetic studies in Arabidopsis in which mto1 mutants, accumulating up to 40-fold higher free Met than wild-type plants, showed DNA sequence alterations within a specific region within this domain, called the MTO1 region (Inba et al, 1994;Chiba et al, 1999). The MTO1 region apparently acts in cis to down-regulate CGS mRNA levels when the level of Met/SAM rises via a mechanism that apparently involves the nascent polypeptide translated from this mRNA region (Chiba et al, 1999;Lambein et al, 2003;Onouchi et al, 2004).…”