Collaborative Research: Manipulating Epigenetic Mechanisms to Enhance Non-Viral Transgene Expression

合作研究：操纵表观遗传机制以增强非病毒转基因表达

• 批准号: 1404084
• 负责人: Kaushal Rege
• 金额: $ 42.64万
• 依托单位: Arizona State University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-01 至 2019-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1404084&HistoricalAwards=false
• 关键词: Collaborative; Research; Manipulating; Epigenetic; Mechanisms

CBET - 1404084/1403214Rege/ElmerDNA or gene delivery to mammalian cells has several applications in biotechnology and medicine. However, the activation of foreign genes (transgenes), and subsequent efficacy of protein expression from them, is often hindered by defense mechanisms inside the host cell. For example, cells are able to control transgene protein expression by either burying existing genes inside condensed chromosomes in the nucleus in order to turn them off, or by relaxing chromosomes in order to expose genes and activate them. A family of 'epigenetic' enzymes present within cells typically controls these processes of gene regulation; enzymes that activate genes are generally classified as activators, while those that turn genes off are called repressors. In the proposed research, the investigators will study the role of these epigenetic enzymes in regulating the expression from delivered transgenes. Furthermore, tools from the emerging field of synthetic biology will be employed to design novel DNA sequences and activator enzymes that enhance protein expression from delivered transgenes. Findings from these studies will have a transformative impact on biotechnology processes and medical treatments (e.g. gene therapy) that utilize DNA or gene delivery to mammalian cells. Outreach from this project will reach K-12 students through a PCR-based molecular biology project facilitated by the Quanta program at ASU. The proposed outreach will be designed to increase participation of students from underrepresented populations in STEM activities in Arizona and Pennsylvania.In the same way that words are neatly organized in the pages of books in a library, cellular genes are condensed by histone proteins to form chromosomes in the nucleus. Epigenetic regulation, mediated by several enzymes, plays a key role in controlling gene expression inside cells. Since recent work has shown that epigenetic mechanisms can silence viral genes, it is likely that epigenetics may also regulate the expression of foreign genes (i.e. transgenes) that are delivered in the form of plasmid DNA to mammalian cells. The goal of this project is to manipulate intracellular epigenetic enzymes in order to enhance transgene expression by (1) using drugs to inhibit repressors and to prevent the deactivation of transgenes, (2) adding designed DNA sequences to the plasmids in order to recruit activator enzymes, and (3) using principles from synthetic biology in order to design and validate novel epigenetic activator enzymes that can selectively bind transgenes and enhance their expression. These approaches will deepen our understanding of epigenetic mechanisms that regulate protein expression from foreign transgenes (plasmid DNA) in human cells, leading to increased efficacy of gene therapy treatments in medicine, and cellular engineering approaches in biotechnology.This award is co-funded by the Biotechnology, Biochemical, and Biomass Engineering Program of the CBET Division and by the Systems and Synthetic Biology Program of the Division of Molecular and Cellular Biology.

CBET -1404084/1403214REGE/ELMERDNA或基因递送到哺乳动物细胞中有多种应用在生物技术和医学中。但是，外源基因（转基因）的激活以及蛋白质表达的随后疗效通常受到宿主细胞内部的防御机制的阻碍。例如，细胞能够通过将现有基因埋在细胞核中的凝结染色体内，以将其关闭，或者通过放松染色体以暴露基因并激活它们来控制转基因蛋白表达。细胞中存在的“表观遗传学”酶家族通常控制这些基因调节过程。激活基因的酶通常被归类为活化剂，而关闭基因的酶称为抑制剂。在拟议的研究中，研究人员将研究这些表观遗传酶在调节传递转基因表达中的作用。此外，将采用合成生物学新兴领域的工具来设计新型的DNA序列和激活酶，从而增强了传递转基因的蛋白质表达。这些研究的发现将对利用DNA或基因递送至哺乳动物细胞的生物技术过程和医疗治疗（例如基因治疗）产生变革性的影响。该项目的推广将通过ASU Quanta计划促进的基于PCR的分子生物学项目吸引K-12学生。拟议的外展活动将旨在增加代表性不足的人群在亚利桑那州和宾夕法尼亚州的STEM活动中的参与。就像单词在图书馆的书籍页面中整齐组织的单词一样，细胞基因在核心中由组蛋白凝结，以形成核心蛋白。由几种酶介导的表观遗传调节在控制细胞内基因表达方面起关键作用。由于最近的工作表明表观遗传机制可以使病毒基因保持沉默，因此表观遗传学可能还可以调节以质粒DNA形式传递给哺乳动物细胞的外生基因（即转基因）的表达。该项目的目的是操纵细胞内表观遗传酶，以通过（1）使用药物来增强转基因表达，以抑制压抑子并防止转基因失活，（2）将设计的DNA序列添加到质粒中，从而将设计的DNA序列添加到质粒中，以便使用募集的活性酶和（3）的启动剂，（3），（3）可以选择性地结合转基因并增强其表达。 These approaches will deepen our understanding of epigenetic mechanisms that regulate protein expression from foreign transgenes (plasmid DNA) in human cells, leading to increased efficacy of gene therapy treatments in medicine, and cellular engineering approaches in biotechnology.This award is co-funded by the Biotechnology, Biochemical, and Biomass Engineering Program of the CBET Division and by the Systems and Synthetic Biology Program of the Division of分子和细胞生物学。