Control of cellular damage: dissecting the physiological functions of the glyoxalase system in plants

反应性羰基物质（RCS）是在细胞代谢过程中形成的高度反应性的小的亲电单和二羰基分子。在人类中，RCS参与几种退行性过程和疾病。在植物中，最丰富的RCS是乙二醛（GO）和甲基乙二醛（MGO），其形成与糖酵解和Calvin-Benson循环的通量密切相关。RCS积累在植物组织暴露于各种非生物胁迫，如盐，干旱和寒冷。这种联系促使人们对通过RCS清除系统提高植物适应性和产量的策略进行了深入研究。GO和MGO解毒的主要途径是谷胱甘肽酶（GLX）系统，该系统由两种酶，S-D-乳酰谷胱甘肽裂解酶（glycoproteinase I，GLXI）和S-2-羟酰基谷胱甘肽水解酶（glycoproteinase II，GLXII），依次作用。植物还具有GLXI样蛋白，其与GLXI蛋白一起属于邻位氧螯合物超家族。我们以前的工作表明，（i）一组特定的GLXI蛋白是唯一的Viridiplantae和可能共同进化的一簇Viridiplantae特异性GLXII蛋白;和（ii）拟南芥GLXI同种型似乎在体内不同的RCS不同的特异性。在这项提案中，我们的目标是澄清参与不同的GLXI和GLXII活动的细胞损伤的控制不同的RCS。为了实现这一目标，我们建议遵循互补的方法，包括（i）具有特定GLXI和GLXII同种型的修饰表达的植物系的功能表征;（ii）重组GLXI的生物化学表征;（iii）使用非靶向代谢组学来鉴定GLXI同种型的其它底物;（iv）使用非靶向代谢组学来鉴定GLXI同种型的其它底物。和（iv）使用互补蛋白质-蛋白质相互作用方法来分析GLXI和GLXII同种型是否物理相互作用以有效地引导反应中间体。

Reactive carbonyl species (RCS) are highly reactive small electrophilic mono- and di-carbonyl molecules formed during cellular metabolism. In humans, RCS are involved in several degenerative processes and diseases. In plants, the most abundant RCS are glyoxal (GO) and methylglyoxal (MGO), whose formation is closely connected to the fluxes through glycolysis and the Calvin-Benson cycle. RCS accumulate in plant tissues upon exposure to various abiotic stresses such as salinity, drought, and cold. This connection has prompted intense research on strategies for the enhancement of plant fitness and yield through RCS scavenging systems. The main pathway to detoxify GO and MGO is the glyoxalase (GLX) system, which consists of two enzymes, S-D-lactoylglutathione lyase (glyoxalase I, GLXI) and S-2-hydroxyacylglutathione hydrolase (glyoxalase II, GLXII), acting in sequence. Plants possess also GLXI-like proteins, which, together with the GLXI proteins, belong to the vicinal oxygen chelate superfamily. Our previous work suggests that (i) a specific group of GLXI proteins are unique to the Viridiplantae and likely co-evolved with a cluster of Viridiplantae-specific GLXII proteins; and (ii) Arabidopsis GLXI isoforms seem to act in vivo on different RCS with different specificity. In this proposal, we aim to clarify the participation of different GLXI and GLXII activities in the control of cellular damage by different RCS. To achieve this aim, we propose to follow complementary approaches, including (i) the functional characterization of plant lines with modified expression of specific GLXI and GLXII isoforms; (ii) the biochemical characterization of recombinant GLXI; (iii) the use of untargeted metabolomics to pursue the identification of additional substrates of the GLXI isoforms; and (iv) to use complementary protein-protein interaction approaches to analyse if GLXI and GLXII isoforms physically interact to efficiently channel the intermediates of the reactions.