Utilizing TALEN technology to regulate human microRNA's

利用TALEN技术调控人类microRNA

• 批准号: 8568549
• 负责人: Ryan M O'Connell
• 金额: $ 223.5万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-30 至 2018-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8568549
• 关键词: Autoimmunity; Binding Sites; Biological Assay; Biology; Cardiovascular Diseases; Code; DNA-Binding Proteins; Disease; Engineering; Gene Deletion; Genes; Germ Lines; Health; Heart Diseases; Human; Immune response; Inflammatory Response; Link; Malignant Neoplasms; Messenger RNA; MicroRNAs; Mus; Nerve Degeneration; Phenotype; Physiology; Proteins; Regulation; Role; Series; Technology; Time; Tumor Suppressor Proteins; Untranslated Regions; combat; homologous recombination; human disease; improved; in vivo; man; novel strategies; nuclease; public health relevance; transcription factor

DESCRIPTION (provided by applicant): MicroRNAs (miRNAs) have quickly emerged as important regulators of mammalian physiology owing to their precise regulation of critical protein coding genes. Importantly, perturbations in expression or function of specific miRNAs has been linked to a plethora of human pathological conditions including cancer, autoimmunity, cardiovascular disease and neurodegeneration. Although miRNA gene deletion in mice via homologous recombination has led to an improved understanding of how they function during times of health and disease, there remains a fundamental need to manipulate miRNA genes and their target binding sites in the human germ line. This would allow for the study of their biology in man, and may be a strategic means by which miRNAs can be targeted therapeutically to combat human disease. We propose to develop a novel approach to activating, repressing or disrupting human miRNA genes in vivo by engineering TALE proteins that will function as transcription factors or nucleases that specifically target miRNA genes or their binding sites in the 3'UTRs of key target mRNAs. We will focus on targeting human miRNA---155, an "oncomiR" involved in regulating immune responses, and miR---146a, a tumor suppressor miRNA that inhibits inflammatory responses. Following proper targeting and disruption of these miRNAs or their binding sites in the 3' UTRs of relevant target mRNAs, a series of functional assays will be carried out to assess the roles of human miR---155 and miR---146a in regulating immune responses and cancer phenotypes. Beyond these two miRNAs, this approach will be compatible with any human miRNA making it a powerful, specific and versatile technology that has not yet been used to study miRNAs.

描述(申请人提供)：由于对关键蛋白质编码基因的精确调控，microRNAs(MiRNAs)迅速成为哺乳动物生理的重要调节因子。重要的是，特定miRNAs表达或功能的紊乱与包括癌症、自身免疫、心血管疾病和神经变性在内的大量人类病理疾病有关。虽然小鼠通过同源重组的miRNA基因缺失导致了对它们在健康和疾病时期如何发挥作用的更好的理解，但仍然有必要操纵miRNA基因及其在人类生殖细胞系中的靶结合位置。这将允许在人类身上研究miRNAs的生物学，并可能成为一种战略手段，通过这种手段，miRNAs可以成为治疗的靶点，以对抗人类疾病。我们建议开发一种新的方法来激活、抑制或破坏体内的人类miRNA基因，方法是设计TALE蛋白作为转录因子或核酸酶，特异性地靶向关键靶向mRNAs的3‘UTRs中的miRNA基因或其结合部位。我们将重点针对人类miRNA-155，一种参与调节免疫反应的“oncomir”，以及miR-146a，一种抑制炎症反应的肿瘤抑制因子。在适当地靶向和破坏这些miRNAs或其在相关靶mRNAs的3‘UTRs中的结合位置后，将进行一系列功能分析来评估人miR--155和miR-146a在调节免疫反应和癌症表型中的作用。除了这两种miRNAs之外，这种方法将与任何人类miRNA兼容，使其成为一种尚未被用于研究miRNAs的强大、特定和通用的技术。