Biosynthesis, Regulation and Engineering of C-Lignin

木质素是地球上最丰富的天然聚合物之一，是木本植物材料的关键组分，也是加工植物材料以生产液体生物燃料的生物精炼厂中的主要废物。首席研究员和合作者发现了一种新型的木质素，显然仅限于一系列非作物物种的种皮。这种新型的木质素具有转化为碳纤维的重大前景。如果这种类型的木质素可以被改造成生物能源原料，它将代表生物精炼的高价值副产品。该项目将确定这种新型木质素合成的生化途径，以及使其在植物茎和叶中积累所需的步骤。了解不同类型的木质素在不同植物组织和细胞类型中积累的限制也解决了细胞生物学中的一个基本问题。该项目采用了一系列方法，从遗传机制到代谢工程，再到生物材料的生产和测试。推广活动将针对两个人口统计学上不同的群体;在数学和科学的得克萨斯州学院，数学和科学的公立学校教师，主要是西班牙裔学生谁是有限的英语熟练（LEP）的高天赋的高中生。教师将开发西班牙语和英语的课程活动，在他们的课堂上实施。该项目将包括非正式的学习活动，通过部署项目，鼓励K-12女孩通过与达拉斯女工程师协会的外联进入STEM。在木质素生物合成过程中，单醇前体通过芳族羟基化和O-甲基化功能化，依次生成羟基苯基（H）、邻苯二酚（C）、愈创木基（G）、5-羟基愈创木基（5-OH-G）和双辛基（S）单元。被子植物中的天然木质素由大约等量的G和S单元组成，具有少于2%的H单元，并且没有C或5-OH-G单元;裸子植物木质素类似，但缺乏S单元。5-OH-G单元被掺入转基因植物中的木质素中，其中第二甲基化步骤被阻断，但尚未报道C-单元在被子植物的木质素中积累，其中第一甲基化步骤类似地被阻断。主要研究人员已经表明，广泛的双子叶植物和单子叶植物的种皮含有一种完全来自儿茶素单元的新型木质素（C-木质素）。这种线性聚合物具有转化为碳纤维的有利性能。然而，C-木质素在自然界中似乎仅限于种皮。 该奖项将解决两个主要问题：植物种皮中C-木质素生物合成的潜在机制是什么，以及为什么C-木质素不天然存在于植物的营养组织中？研究人员假设，C-木质素生物合成涉及经典的单体生物合成中的一个或两个甲基化步骤的功能丧失，并将使用转录组分析来确定这种情况如何发生在Cleome hassleriana的发育种子中，其中在授粉后约13天，从G木质素到C木质素生物合成的突然转变。解决第二个问题将涉及尝试引入C-木质素聚合物到不同组织的模式豆科苜蓿，利用现有的突变体，其中木质素甲基化酶已被功能性破坏。 这些研究旨在确定C-木质素在营养组织中以足够的产量进行工程积累以实现工业加工的潜力，并且在这种情况下，将开发用于从种子和营养组织中提取C-木质素的改进方法，并进一步分析聚合物的物理性质。

Lignin is one of earth's most abundant natural polymers, a key component of woody plant material, and a major waste product in bio-refineries that process plant material for production of liquid biofuels. The lead investigator and collaborators have discovered a new type of lignin, apparently restricted to the seed coats of a range of non-crop species. This new type of lignin has significant promise for conversion to carbon fibers. If this type of lignin could be engineered into bioenergy feedstocks, it would represent a high value co-product of biorefining. This project will determine the biochemical pathway by which this new type of lignin is synthesized, and the steps that will need to be taken to make it accumulate in the stems and leaves of plants. Understanding the limitations to the accumulation of different types of lignin in different plant tissue and cell types also addresses a fundamental problem in cell biology. This project uses a range of approaches from genetic mechanisms through metabolic engineering to production and testing of biomaterials. Outreach activities will target two demographically distinct groups; highly talented high school students at the Texas Academy of Mathematics and Science, and mathematics and science public school teachers with primarily Hispanic students who are Limited English Proficient (LEP). Teachers will develop curricular activities in Spanish and English to implement in their classrooms. The project will include informal learning activities through deployment of the projects to encourage K-12 girls into STEM through outreach with the Dallas Society of Women Engineers.During lignin biosynthesis, the monolignol precursors are functionalized by aromatic hydroxylation and O-methylation to generate, successively, hydroxyphenyl (H), catechyl (C), guaiacyl (G), 5-hydroxyguaiacyl (5-OH-G), and syringyl (S) units. Natural lignins in angiosperms consist of approximately equal amounts of G and S units, with less than 2% of H units and no C or 5-OH-G units; gymnosperm lignins are similar but lack S units. 5-OH-G units are incorporated into lignins in transgenic plants in which the second methylation step is blocked, but C-units have not been reported to accumulate in the lignin of angiosperms in which the first methylation step is similarly blocked. The Principal Investigators have shown that the seed coats of a wide range of dicot and monocot plants contain a novel type of lignin (C-lignin) derived totally from catechyl units. This linear polymer has advantageous properties for conversion to carbon fibers. However, C-lignin appears to be restricted to seed coats in nature. This award will address two major questions: what is the mechanism underlying the biosynthesis of C-lignin in plant seed coats, and why does C-lignin not occur naturally in the vegetative tissues of the plant? The researchers hypothesize that C-lignin biosynthesis involves loss of function of one or both methylation steps in classical monolignol biosynthesis, and will determine how this happens using transcriptome profiling in developing seeds of Cleome hassleriana, in which there is an abrupt transition from G lignin to C lignin biosynthesis at around 13 days post-pollination. Addressing the second question will involve attempting to introduce the C-lignin polymer into different tissues of the model legume Medicago truncatula, utilizing existing mutants in which enzymes of lignin methylation have been functionally disrupted. These studies aim to determine the potential for engineering accumulation of C-lignin in vegetative tissues in sufficient yields to enable industrial processing, and, in this context, improved methods for extraction of C-lignin from both seeds and vegetative tissues will be developed, and the physical properties of the polymer further analyzed.