ELP

引言紫杉醇是一种众所周知的抗癌化合物。它的生物合成包括形成一个高度功能化的二萜核心骨架(baccatin III)和随后的苯基异丝氨酸侧链的组装。尽管经过半个世纪的深入研究，baccatin III的完整生物合成途径仍然未知。2024年1月25日，中国农业科学院农业基因组研究所闫建斌及北京大学雷晓光共同通讯在Science 在线发表题为”Characterization and heterologous reconstitution of Taxus biosynthetic enzymes leading to baccatin III“的研究论文，该研究发现了一种双功能细胞色素P450酶(Taxane oxetanase, TOT)，它催化紫杉醇oxetane形成中的氧化重排，代表了以前未知的oxetane环形成的酶机制。该研究建立了基于紫杉素（taxusin）生物合成途径的筛选策略，并发现了负责紫杉烷氧化C9位置(T9αH)的酶。最后，该研究在烟草中人工重组了一种生产baccatin III的生物合成途径。总之，该研究成功鉴定了紫杉醇生物途径的关键缺失酶，揭示了红豆杉催化含氧四元环分子形成的全新机制，发现了紫杉醇异源生物合成的核心基因，实现了紫杉醇核心前体baccatin III在烟草中的异源合成。该研究成果解决了紫杉醇生物合成研究中的关键瓶颈问题，是植物天然产物生物合成领域中的重大突破，为通过合成生物学的手段实现紫杉醇的高效、可持续生产铺平了道路，具有极高的科学创新性与应用价值。紫杉醇来源于红豆杉科红豆杉属植物的次生代谢，已被临床用于治疗多种癌症。紫杉醇含有结构复杂的6-8-6三环碳骨架，包含9个立体中心，1个显著的氧烷环基序和1个苯异丝氨酸链。紫杉醇可以穿过微管壁的纳米孔，与微管管腔表面的微管蛋白相互作用，破坏微管动力学，从而触发对癌细胞的细胞毒性作用。三环碳骨架的整体构象硬化是由紫杉醇的氧烷环产生的。目前用于生产紫杉醇的一种策略是化学半合成，即从植物细胞培养物或红豆杉的原始树木生物量中分离出紫杉醇前体，如baccatin III，然后化学转化为紫杉醇。这种半合成策略严重依赖自然资源和红豆杉植物的人工种植，无法满足日益增长的市场需求。以绿色和可持续的方式生产紫杉醇的生物合成策略已经变得非常有吸引力。在红豆杉植物中，TOT1负责氧乙烷的形成（Credit: Science ）自1971年首次发现紫杉醇结构以来，阐明其生物合成途径一直是天然产物研究中的一个具有挑战性的任务。迄今为止，大约有20种可能的酶促反应被报道。反应步骤可分为三个关键过程，包括从底物香叶基焦磷酸(GGPP)形成4(20)-烯-5α-醇片段的紫杉烷骨架，通过C4和C20双键的环氧化生物合成baccatin III，以及将苯基异丝氨酸链连接到baccatin III的C13位置生成紫杉醇。C13侧链的形成及其与baccatin III结合生成紫杉醇的研究已经得到了很好的研究。然而，形成baccatin III的几个基本步骤仍然未知，特别是氧烷环的形成和C9的氧化。因此，紫杉醇生物合成的上游和下游步骤无法连接，迄今为止缺乏完整的紫杉醇生物合成途径。该研究发现了一种双功能细胞色素P450酶(Taxane oxetanase, TOT)，它催化紫杉醇oxetane形成中的氧化重排，代表了以前未知的oxetane环形成的酶机制。该研究建立了基于紫杉素（taxusin）生物合成途径的筛选策略，并发现了负责紫杉烷氧化C9位置(T9αH)的酶。最后，该研究在烟草中人工重组了一种生产baccatin III的生物合成途径。5α-乙酰基4,20烯在TOT1介导的氧乙烷生成反应中是不可缺少的（Credit: Science ）原文链接https://www.science.org/doi/10.1126/science.adj3484责编|探索君排版|探索君文章来源|“iNature”End往期精选围观Nat Med | 黄荷凤院士/张静澜/徐晨明团队发表综合性无创产前筛查临床研究成果热文Science最新发布：全世界最前沿的125个科学问题热文Nature | 2024年值得关注的七项技术热文Let-7家族miRNA或为癌症潜在免疫治疗靶点热文Nat Methods | 自主生物发光成像技术在真核生物中的创新进展

Tweaks to the mitochondrial genome hold the potential for better hybrid seed production or to introduce seedless fruits. North Carolina State University researchers have successfully transferred an important gene from one compartment of a plant cell to another to produce tobacco plants that lack pollen and viable seeds, while otherwise growing normally. Their findings could lead to better ways of producing hybrid seeds to maximize crop productivity, or to introduce seedlessness in fruit species lacking the often-desired trait, such as raspberries, blackberries or muscadine grapes. The researchers began the work in the energy-producing portion of a cell, the mitochondria. In plants, aberrations within the mitochondrial genome can be associated with the inability to make pollen, a trait known as cytoplasmic male sterility (CMS) that has been successfully exploited for the production of high-yielding hybrid seeds in many important crops. Naturally occurring CMS-based systems that are robust enough to facilitate commercial scale hybrid seed production are limited, however. In their proof-of-concept study, the NC State researchers, together with colleagues from Precision BioSciences and Elo Life Systems, deployed a unique strategy to test whether the CMS trait could be generated in tobacco, a commonly used model species in plant research. The researchers initially took an essential mitochondrial gene called atp1 and moved it to the nucleus after placing it under the regulatory control of an element -- known as a promoter -- that they predicted would allow the transferred atp1 gene to be expressed in every cell of the plant except those responsible for producing pollen. The researchers then used genome editing tools to permanently remove the native atp1 gene from the mitochondria. Their approach was successful. "The results exceeded our expectations," said Ralph Dewey, Philip Morris Professor of Crop Science at NC State and corresponding author of a paper describing the research. "The plants looked completely normal up until they began to flower, but then failed to produce pollen because the transferred atp1 gene was no longer expressed. Importantly, because the original atp1 gene was deleted from the mitochondrial genome, the trait will be inherited maternally, which is a crucial consideration for large-scale hybrid seed production." Pollen wasn't the only casualty of the technique. When cross-fertilized using pollen from a neighboring normal plant, their tobacco plants unexpectedly produced small, hollow seeds, much like those seen in popular "seedless" fruits such as watermelons and grapes. "That's because the promoter that we chose not only failed to express during pollen formation, but also during early seed development," Dewey said. Dewey said that his team is now working to decouple these results so that researchers can achieve either pollen infertility or the seedless trait alone, instead of both at the same time. Dewey also stressed that the findings shouldn't be limited to tobacco plants. Their next generation of experiments will include testing the seedless trait in tomato, a close relative of tobacco. They will also test their novel CMS trait in a grain such as rice to test the efficacy of their system in a crop where hybrid seed production is important for achieving maximal yields. "Knowing the way the system works, there's no reason to believe that we couldn't effectively transfer the technology to other plant species," he said. The study appears in Frontiers in Plant Science. Devarshi Selote, H. Carol Griffin, Allison N. Dickey, Derek Jantz, J. Jeff Smith, Anna Matthiadis, Josh Strable, Caitlin Kestell and William A. Smith co-authored the paper. The work was funded in part by a grant from Elo Life Systems and supported by the NC State Plant Breeding Consortium. Dewey and several other researchers have filed a patent on the new technique.

Biologists demonstrate how the auxiliary factor CGL160 contributes to the synthesis of crucial parts of the photosynthetic machinery. LMU biologists demonstrate how the auxiliary factor CGL160 contributes to the synthesis of crucial parts of the photosynthetic machinery. Photosynthesis is the means by which plants, algae, and cyanobacteria extract their 'food' in the form of energy-rich biomolecules from sunlight, carbon dioxide, and water. It is a complex process, from which researchers are still coaxing many new details. A team led by LMU biologists Thilo Rühle, Bennet Reiter, and Prof. Dario Leister has now solved another piece of the puzzle of this essential process and elucidated the role of the auxiliary factor CGL160, as the scientists report in the journal The Plant Cell. Photosynthesis takes place in several partial reactions at the so-called thylakoid membranes in the chloroplasts, where various pigments absorb the electromagnetic radiation of sunlight. Specific protein complexes in conjunction with ATP synthases then convert this light energy into chemical energy in the form of ATP. This enables the plant, among other things, to synthesize carbohydrates as energy-rich 'fuel' for cellular respiration. As 'molecular machines,' ATP synthases are therefore a vital component of plant metabolism. How ATP synthases are assembled in the cell has not yet been fully clarified. Based on the model organism Arabidopsis thaliana, the researchers have now been able to demonstrate that the protein CGL160 plays a key role in the process by recruiting the coupling factor CF1 of the ATP synthase. "The protein CGL160 sits with its base in the thylakoid membrane, while its N-terminal domain protrudes like a rod and fishes the soluble CF1 portion of the ATP synthases out of the fluid inside the chloroplasts. This portion of the protein binds the CF1 'headpiece' and facilitates linkage with the portion of the ATP synthase embedded in the thylakoid membrane, making its formation considerably more efficient," explains Thilo Rühle. Evolutionary roots and adaptations Even under unfavorable growth conditions, the presence of auxiliary factors such as CGL160 prove to be advantageous: The LMU biologists discovered that an absence of CGL160, with its CF1-binding function, under conditions of light deficiency negatively impacts the development of chloroplasts. "We observed that the chloroplast structure of corresponding mutants suffers more during short days with just eight hours of daylight. We advance the hypothesis therefore that the interaction between the N-terminal domain of CGL160 and CF1 is also an evolutionary adaptation that enables plants to cope better in various light conditions," says Rühle. The researchers also found clues as to the evolutionary origin of CGL160. In experiments with Arabidopsis thaliana, they succeeded in functionally replacing the membrane domain of CGL160 with the Atp1 protein from cyanobacteria of the genus Synechocystis. "Around 80 percent of the function can be restored in this way. This suggests that CGL160 in land plants may have come from a predecessor protein of cyanobacteria," says Rühle. Indeed, plant cells acquired their chloroplasts through incorporation and functional integration of cyanobacteria. Potential route to regulation of photosynthesis This set in train the emergence of a complex synthesis plan for photosynthetic machinery. To better understand this plan, the LMU biologists now want to identify all auxiliary factors -- in addition to CGL160 and a handful of other currently known factors -- that are important for the assembly and function of ATP synthases in plant thylakoid membranes. "We assume that there are much fewer factors involved here than in the photosystems themselves -- accordingly, ATP synthase is an easier potential route for the regulation of photosynthesis," says Rühle. "In the future, people could use this knowledge for various applications, such as modifying plants so that they can optimally cover their ATP requirements even when exposed to stressors like heat or extreme light conditions."