Unraveling the biological roles of specific miRNAs, from experimental target identification through functional characterization
从实验目标识别到功能表征,揭示特定 miRNA 的生物学作用
基本信息
- 批准号:10566442
- 负责人:
- 金额:$ 31.67万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2023
- 资助国家:美国
- 起止时间:2023-03-10 至 2027-01-31
- 项目状态:未结题
- 来源:
- 关键词:BindingBiologicalBiological AssayBiological ModelsBiological ProcessCell Differentiation processCell Fate ControlCell LineCell ProliferationCell physiologyCellsComplexDataDiseaseDown-RegulationEmbryonic DevelopmentGene ExpressionGenesGoalsIndividualInjuryKnock-outKnowledgeMediatingMessenger RNAMethodsMicroRNAsModelingMolecularMusMuscle FibersMyoblastsPathologicPathway interactionsPhenotypePlayProcessProliferatingRNARegulationRegulator GenesResearchRoleTechniquesTestingUndifferentiatedValidationWorkbiological systemsexperimental studyin vivoin vivo ModelmRNA Expressionmouse modelmuscle regenerationmyogenesisprematurepreventskeletal muscle differentiationsuccess
项目摘要
PROJECT SUMMARY/ABSTRACT
Precisely controlling the expression levels of genes is essential to any biological process. microRNAs
(miRNAs) play important roles in controlling gene expression by binding to specific target mRNAs and
downregulating their expression. To understand how a miRNA functions it is critical to both identify the set of
mRNA targets to which it binds in a specific biological context, and to determine how this pairing interaction
regulates gene expression to ultimately impact cellular processes and phenotypes. Some miRNAs can be
controlled by competing endogenous RNAs (ceRNAs), which bind miRNAs and prevent them from
downregulating expression of their mRNA targets, creating a complex network of regulation that can be difficult
to experimentally unravel.
The proposed research will develop a comprehensive experimental approach to uncover the biological
functions of a miRNA. As a model system the work focuses on two miRNAs (miR-1 and miR-206) that control
myogenesis, the differentiation of skeletal muscle cells. The experiments use cultured mouse C2C12 cells, a
widely used model for myogenesis, as well as in vivo experiments with mice. In Specific Aim 1 experiments will
identify the direct RNA targets of miR-1 and miR-206 in undifferentiated C2C12 cells and during differentiation.
Supporting data demonstrate the success of a target identification technique, which will be used with miR-1
and miR-206 knockout cells to identify mRNAs uniquely targeted by each miRNA. In Specific Aim 2, newly
identified targets of miR-1 and miR-206 during myogenesis will be screened for their contributions to
differentiation phenotypes. Experiments will also test the function of miRNA/target pairing in controlling the
expression levels of targets during myogenesis. In addition, the regulatory relationships of specific miRNA/
target pairs will be studied using mouse models. In Specific Aim 3 experiments will test the hypothesis that
RNA targets of miR-1 and miR-206 in undifferentiated cells function as ceRNAs to maintain proliferation and
prevent premature differentiation. The absolute levels of miRNAs and their targets in undifferentiated cells will
be quantified, targets will be screened to identify those important for maintaining cellular proliferation, and
regulatory mechanisms of ceRNAs will be investigated.
Together the proposed studies develop an experimental framework to understand the functional roles of
two important miRNAs. This work will not only advance understanding of the roles of miR-1 and miR-206 in
myogenesis, but will also provide a comprehensive strategy that could be applied to other miRNAs in a variety
of biological systems.
项目总结/摘要
精确控制基因的表达水平对任何生物过程都是必不可少的。微rna
(miRNAs)通过与特定的靶mRNA结合,在控制基因表达方面发挥重要作用,
下调它们的表达。为了了解miRNA的功能,关键是要识别一组
mRNA在特定的生物学背景下结合的靶点,并确定这种配对相互作用
调节基因表达以最终影响细胞过程和表型。一些miRNAs可以
由竞争性内源性RNA(ceRNA)控制,其结合miRNA并阻止它们
下调其mRNA靶点的表达,形成一个复杂的调控网络,
实验性地解开。
拟议的研究将开发一种全面的实验方法,以揭示生物学
miRNA的功能。作为一个模型系统,这项工作集中在两个miRNAs(miR-1和miR-206),
骨骼肌细胞的分化。实验使用培养的小鼠C2 C12细胞,
广泛用于肌生成的模型,以及小鼠体内实验。在具体目标1实验中,
在未分化的C2 C12细胞中和分化期间鉴定miR-1和miR-206的直接RNA靶。
支持性数据证明了将用于miR-1的靶点鉴定技术的成功
和miR-206敲除细胞以鉴定每种miRNA独特靶向的mRNA。在具体目标2中,新
将筛选肌生成过程中miR-1和miR-206的已鉴定艾德靶点,以确定它们对肌生成的贡献。
分化表型实验还将测试miRNA/靶标配对在控制细胞凋亡中的功能。
在肌生成过程中的目标的表达水平。此外,特定miRNA/的调节关系
将使用小鼠模型研究靶对。在具体目标3中,实验将检验以下假设:
未分化细胞中miR-1和miR-206的RNA靶标作为ceRNA起作用以维持增殖和分化。
防止过早分化。未分化细胞中miRNAs及其靶标的绝对水平将
艾德,筛选靶点以确定对维持细胞增殖重要的靶点,
将研究ceRNA的调节机制。
这些拟议的研究共同建立了一个实验框架,以了解
两个重要的miRNAs这项工作不仅将促进对miR-1和miR-206在细胞凋亡中作用的理解,
肌生成,但也将提供一个全面的战略,可适用于其他miRNAs在各种
生物系统。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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Jennifer F. Kugel其他文献
Non-coding-RNA regulators of RNA polymerase II transcription
RNA 聚合酶 II 转录的非编码 RNA 调节因子
- DOI:
10.1038/nrm1946 - 发表时间:
2006-05-24 - 期刊:
- 影响因子:90.200
- 作者:
James A. Goodrich;Jennifer F. Kugel - 通讯作者:
Jennifer F. Kugel
Correction for Abrisch et al., Infection by Herpes Simplex Virus 1 Causes Near-Complete Loss of RNA Polymerase II Occupancy on the Host Cell Genome
更正 Abrisch 等人,单纯疱疹病毒 1 感染导致宿主细胞基因组上 RNA 聚合酶 II 占据几乎完全丧失
- DOI:
- 发表时间:
2016 - 期刊:
- 影响因子:5.4
- 作者:
Robert G. Abrisch;Tess M. Eidem;Petro Yakovchuk;Jennifer F. Kugel;J. Goodrich - 通讯作者:
J. Goodrich
Release of human TFIIB from actively transcribing complexes is triggered upon synthesis of 7 nt and 9 nt RNAs
7 nt 和 9 nt RNA 合成后,触发主动转录复合物释放人 TFIIB
- DOI:
- 发表时间:
2019 - 期刊:
- 影响因子:0
- 作者:
Elina Ly;Abigail E Powell;J. Goodrich;Jennifer F. Kugel - 通讯作者:
Jennifer F. Kugel
In vitro studies of the early steps of RNA synthesis by human RNA polymerase II.
人 RNA 聚合酶 II 合成 RNA 早期步骤的体外研究。
- DOI:
10.1016/s0076-6879(03)70056-1 - 发表时间:
2003 - 期刊:
- 影响因子:0
- 作者:
Jennifer F. Kugel;J. Goodrich - 通讯作者:
J. Goodrich
Single molecule FRET shows uniformity in TBP-induced DNA bending and heterogeneity in bending kinetics †
单分子 FRET 显示 TBP 诱导的 DNA 弯曲的均匀性和弯曲动力学的异质性 †
- DOI:
- 发表时间:
2013 - 期刊:
- 影响因子:0
- 作者:
Rebecca H. Blair;J. Goodrich;Jennifer F. Kugel - 通讯作者:
Jennifer F. Kugel
Jennifer F. Kugel的其他文献
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{{ truncateString('Jennifer F. Kugel', 18)}}的其他基金
Defining the Genome-wide Alcohol-induced Transcriptional Changes in Breast Cancer
定义酒精诱导的乳腺癌全基因组转录变化
- 批准号:
9761407 - 财政年份:2018
- 资助金额:
$ 31.67万 - 项目类别:
Identify the Transcriptome and Proteome Associated with miRNAs dring Myogenesis
鉴定与肌发生过程中 miRNA 相关的转录组和蛋白质组
- 批准号:
8866691 - 财政年份:2015
- 资助金额:
$ 31.67万 - 项目类别:
Identify the Transcriptome and Proteome Associated with miRNAs dring Myogenesis
鉴定与肌发生过程中 miRNA 相关的转录组和蛋白质组
- 批准号:
9038986 - 财政年份:2015
- 资助金额:
$ 31.67万 - 项目类别:
Controlling NFAT1 in T cells using engineered ncRNA transcriptional regulators
使用工程化 ncRNA 转录调节因子控制 T 细胞中的 NFAT1
- 批准号:
7509919 - 财政年份:2008
- 资助金额:
$ 31.67万 - 项目类别:
Controlling NFAT1 in T cells using engineered ncRNA transcriptional regulators
使用工程化 ncRNA 转录调节因子控制 T 细胞中的 NFAT1
- 批准号:
7632171 - 财政年份:2008
- 资助金额:
$ 31.67万 - 项目类别:
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