Molecular mechanism of translation arrest and mRNA degradation in cystathionine gamma-synthase, the key-step enzyme of methionine biosynthesis.

胱硫醚γ-合酶（甲硫氨酸生物合成的关键步骤酶）翻译停滞和 mRNA 降解的分子机制。

• 批准号: 16370016
• 负责人: NAITO Satoshi
• 金额: $ 9.86万
• 依托单位: HOKKAIDO UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2004
• 资助国家: 日本
• 起止时间: 2004 至 2006
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-16370016/
• 关键词: Metabolic regulation; Cystathionine gamma-synthase; S-Adenosylmethionine; Methionine; mRNA degradation; Arabidopsis; Mutations; シスタチオニン γ-シンターゼ; 突然変異株; 翻訳終止コドン

Cystathionine gamma-synthase (CGS) of Arabidopsis catalyzes the key step in methionine biosynthesis. Unlike many of the key-step enzymes in biosynthetic pathways, CGS is not an allosteric enzyme. Expression of CGS is feedback-regulated at the step of mRNA degradation and S-adenosylmethionine (SAM) acts as the effector. Upon mRNA degradation, 5'-truncated RNA species are formed as degradation intermediates. A short stretch of amino acids, termed the MTO1 region, located within the first exon-coding region of CGS itself, functions as the cis element in this regulation. CGS regulation occurs during translation and is reproduced in the in vitro translation system of wheat germ extract. Studies using the in vitro system revealed that, prior to the mRNA degradation, SAM induces translation elongation arrest at around the MTO1 region. Thus, translation elongation arrest triggers the mRNA degradation. Finer mapping of the translation arrest site showed that translation arrest occurs at Ser-94 located right after the MTO1 region, and peptidyl-tRNA(Ser) accumulates. Alanine substitution experiments identified that, in addition to the MTO1 region amino acid sequence, Trp-93 and Ser-94 are important for the translation arrest. A nascent peptide emerges from the translating ribosome through the ribosomal exit tunnel. This exit tunnel is 15-20 Angstroms in diameter and about 100 Angstroms long, and 30-40 amino acid residues of the nascent peptide will be located in the tunnel. Relative positions of the MTO1 region and the arrest site (Ser-94) suggest that the MTO1 region nascent peptide is located within the exit tunnel. For genetic analyses of the CGS regulation, Arabidopsis mutants that alter the post-transcriptional regulation of CGS expression were isolated and their responsive genes have been identified.

拟南芥的胱硫醚-γ-合成酶(CGS)催化甲硫氨酸生物合成的关键步骤。与生物合成途径中的许多关键步骤酶不同，CGS不是变构酶。Cgs的表达在基因的降解过程中受到反馈调节，S-腺苷蛋氨酸作为效应因子。在信使核糖核酸降解时，形成5‘-截短的RNA物种作为降解中间产物。位于CGS自身第一外显子编码区的一小段氨基酸称为MTO1区，在这一调控中起顺式元件的作用。CGS调控发生在翻译过程中，并在小麦胚芽提取物的体外翻译系统中复制。使用体外系统的研究表明，在mRNA降解之前，SAM在MTO1区域周围诱导翻译延伸停滞。因此，翻译延长停滞触发了mRNA的降解。对翻译抑制位点的精细定位表明，翻译抑制发生在紧挨着MTO1区域的Ser-94处，并积聚了多肽-tRNA(Ser)。丙氨酸替代实验证明，除了MTO1区的氨基酸序列外，Trp-93和Ser-94对翻译抑制也是重要的。翻译的核糖体通过核糖体出口通道产生新生多肽。这个出口通道的直径为15-20埃，长约100埃，新生肽的30-40个氨基酸残基将位于隧道内。MTO1区和停滞部位(Ser-94)的相对位置表明，MTO1区新生多肽位于出口隧道内。为了对CGS的调控进行遗传分析，我们分离了改变CGS表达转录后调控的拟南芥突变体，并鉴定了它们的反应基因。

项目成果

Nascent peptide-mediated translation elongation arrest coupled with mRNA degradation in the CGS1 gene of Arabidopsis.

拟南芥 CGS1 基因中新生肽介导的翻译延伸停滞与 mRNA 降解相结合。

• 发表时间: 2005
• 期刊: Genes Dev. 19
• 作者: Tetsuhiro Ogawa;Sakura Inoue;Shunsuke Yajima;Makoto Hidaka;Haruhiko Masaki;K.Takahashi;S.Ohashi-Kumihiro;T. Takayama;H. Onouchi;T. Takayama;H. Onouchi
• 通讯作者: H. Onouchi

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

• DOI: 10.1042/bst0320597
• 发表时间: 2004-08-01
• 期刊: BIOCHEMICAL SOCIETY TRANSACTIONS
• 影响因子: 3.9
• 作者: Onouchi, H;Lambein, I;Naito, S
• 通讯作者: Naito, S

Proteomic and transcriptomic analysis of Arabidopsis seeds: molecular evidence for successive processing of seed proteins and its implication in the stress response to sulfur nutrition

• DOI: 10.1111/j.1365-313x.2006.02900.x
• 发表时间: 2006-11-01
• 期刊: PLANT JOURNAL
• 影响因子: 7.2
• 作者: Higashi, Yasuhiro;Hirai, Masami Yokota;Saito, Kazuki
• 通讯作者: Saito, Kazuki

Nascent peptide-mediated translation elongation arrest coupled with mRNA degradation in the CGS gene of Arabidopsis.

拟南芥 CGS 基因中新生肽介导的翻译延伸停滞与 mRNA 降解相结合。

• 发表时间: 2005
• 期刊: Genes Dev. 19
• 作者: Onouchi H;Nagami Y;Haraguchi Y;Nakamoto M;Nishimura Y;Sakurai R;Nagao N;Kawasaki D;Kadokura Y;Naito S
• 通讯作者: Naito S