The Impact of Sterol Biogenesis on Cellulose Synthesis in Higher Plants

纤维素是植物细胞壁的结构成分，是地球上最丰富的天然聚合物。它是由纤维素合酶产生的，纤维素合酶是一种存在于质膜上的大型多亚基复合物。利用拟南芥模型进行的遗传研究已经鉴定出了纤维素合酶机制的蛋白质，以及与核心复合物相互作用的关键酶和细胞骨架蛋白。膜脂或固醇等其他因素的潜在作用是有趣的，但尚未充分探索。先前的研究表明，甾醇成分对纤维素合酶活性至关重要。然而，甾醇影响纤维素合成的分子机制至今尚不清楚。在这个项目中，将测试两个有争议的假设，以解释甾醇和纤维素合成之间的联系。首先，甾醇糖苷是植物中丰富的一类甾醇缀合物，通过将葡萄糖残基转移到纤维素微原纤维的生长链中，作为合成的引物。另外，甾醇在纤维素合成中扮演着重要的角色，是抗洗涤剂质膜子域的组成部分。甾醇可以维持跨膜蛋白的结构和完整性，例如组成纤维素合酶的亚基。为了确定哪些机制发生在细胞中，该项目将采用跨学科的方法，以初级细胞壁为模型。将应用遗传学和化学遗传学，生物化学和细胞生物学，以及跨越基因组学，脂质组学和蛋白质组学的系统生物学方法。甾醇生物合成途径中酶的化学抑制剂和拟南芥的突变体将被用作剖析纤维素合酶功能对膜的要求的工具。本项目的最终目标是阐明甾醇和纤维素合成在构建细胞壁中的分子联系。这项研究有望在理解纤维素合成方面取得变革性进展，并有很强的潜力揭示植物细胞质膜中固醇和蛋白质之间动态相互作用的新见解。更广泛的影响：本研究将通过博士后、研究生和本科生的多学科教育产生更广泛的影响。代表性不足的群体将通过麦克奈尔学者、暑期本科生研究机会和路易斯·斯托克斯少数民族参与联盟等项目招募。通过与当地教师合作，以及为对科学、技术、工程和数学职业感兴趣的女孩开设的项目，将实现K-12的推广。通过与瑞典一个实验室的国际合作，该项目对培养具有全球视野的下一代美国科学家产生了更广泛的影响。从这项研究中获得的信息将有助于改进植物衍生产品和生物技术中与生物质有关的应用。破译膜环境如何影响纤维素合酶机制可能会导致有关纤维素的消化率和细胞壁中产生的纤维素量的知识，这是未来工程生物能源作物的关键考虑因素。

Cellulose is the structural component of the plant cell wall and the most abundant natural polymer on earth. It is produced by cellulose synthase, a large multi-subunit complex that resides in the plasma membrane. Genetic studies using the model Arabidopsis have led to the identification of proteins comprising the cellulose synthase machinery as well as critical enzymes and cytoskeletal proteins that interact with the core complex. A potential role for additional factors such as membrane lipids or sterols is intriguing but has not yet been adequately explored. Previous studies demonstrated that sterol composition is crucial for cellulose synthase activity. Yet, the molecular mechanisms by which sterols influence cellulose synthesis are not understood to date. In this project two debated hypotheses to explain the connection between sterols and cellulose synthesis will be tested. The first is that steryl glucosides, an abundant class of sterol conjugates in plants, act as primers for synthesis by transferring glucose residues to the growing chain of cellulose microfibrils. Alternatively or in addition, sterols play a general role in cellulose synthesis, as integral components of detergent-resistant plasma membrane subdomains. Sterols may act to maintain the structure and integrity of transmembrane proteins, such as the subunits that comprise cellulose synthase. To determine which of these mechanisms occurs in the cell, the project will employ an interdisciplinary approach using the primary cell wall as a model. Genetics and chemical genetics, biochemistry and cell biology, in parallel with systems biology approaches spanning genomics, lipidomics and proteomics will be applied. Chemical inhibitors of enzymes in the sterol biosynthesis pathway and mutants of Arabidopsis will be used as tools to dissect the membrane requirements for cellulose synthase function. The ultimate goal of this project is to elucidate the molecular link between sterols and cellulose synthesis in building the cell wall. This research is expected to result in transformative advances in understanding cellulose synthesis, and has strong potential to uncover novel insights in the dynamic interplay between sterols and proteins in the plasma membrane of plant cells. Broader Impacts: This research will have broader impacts through the multi-disciplinary education of a postdoctoral scholar, graduate and undergraduate students. Underrepresented groups will be recruited through programs for McNair Scholars, Summer Undergraduate Research Opportunity, and Louis Stokes Alliance for Minority Participation. K-12 outreach will be achieved by partnerships with local teachers and programs for girls with interests in science, technology, engineering and mathematics careers. Through an international collaboration with a laboratory in Sweden, this project has broader impacts in training the next generation of US scientists with a global perspective. The information gained from this research will facilitate improvements in plant-derived products and biomass-related applications in biotechnology. Deciphering how the membrane environment influences cellulose synthase machinery may lead to knowledge concerning the digestibility of cellulose and the amount of cellulose that is produced in the cell wall, which are key considerations in engineering bioenergy crops of the future.