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，也是地球上含量最丰富的天然聚合物。它是由纤维素合酶产生的，这是一种存在于质膜中的大型多亚单位复合体。利用拟南芥模型进行的遗传学研究已经确定了包含纤维素合成酶机制的蛋白质以及与核心复合体相互作用的关键酶和细胞骨架蛋白。膜脂或甾醇等其他因素的潜在作用是耐人寻味的，但尚未得到充分研究。以往的研究表明，甾醇的组成对纤维素合酶的活性至关重要。然而，迄今为止，甾醇影响纤维素合成的分子机制尚不清楚。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。第一种是甾基葡萄糖苷，这是植物中一类丰富的甾醇偶联物，通过将葡萄糖残留物转移到不断生长的纤维素微纤维链上，作为合成的底物。作为替代或补充，甾醇作为耐洗涤剂的质膜亚区的组成部分，在纤维素合成中起着普遍的作用。甾醇可能起到维持跨膜蛋白的结构和完整性的作用，例如组成纤维素合成酶的亚单位。为了确定这些机制中的哪一种发生在细胞中，该项目将使用以初级细胞壁为模型的跨学科方法。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、脂类组学和蛋白质组学的系统生物学方法将被应用。类固醇生物合成途径中的酶的化学抑制剂和拟南芥的突变体将被用作分析纤维素合成酶功能所需的膜的工具。这个项目的最终目标是阐明在构建细胞壁的过程中，甾醇和纤维素合成之间的分子联系。这项研究有望在理解纤维素合成方面取得革命性的进展，并有很强的潜力揭示植物细胞质膜中甾醇和蛋白质之间的动态相互作用。更广泛的影响：这项研究将通过对博士后学者、研究生和本科生的多学科教育产生更广泛的影响。代表不足的群体将通过麦克奈尔学者计划、夏季本科生研究机会计划和路易斯·斯托克斯少数群体参与联盟计划招募。将通过与当地教师的伙伴关系以及针对对科学、技术、工程和数学职业感兴趣的女孩的项目来实现K-12的推广。通过与瑞典实验室的国际合作，该项目在培养具有全球视野的下一代美国科学家方面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量有关的应用。破译膜环境是如何影响纤维素合成酶机制的，可能会带来关于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.