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Mechanisms in transcriptional regulation during cardiac hypertrophy

心脏肥大过程中的转录调控机制

基本信息

批准号：
9064832
负责人：
Maha Abdellatif
金额：
$ 39.75万
依托单位：
RBHS-NEW JERSEY MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-08-26 至 2018-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9064832
关键词：
Actins Attention CDK9 Protein Kinase Cardiac Cardiac Myocytes Cardiomyopathies Cells Collagen DNA Data Development Disease model Down-Regulation Drug Design Essential Genes Extracellular Matrix Functional disorder Future Gene Expression Gene Expression Regulation Genes Genetic Transcription Grant Health Heart Heart Hypertrophy Heart failure Housekeeping Housekeeping Gene Hypertrophy Immune response Knowledge MAPK1 gene Messenger RNA MicroRNAs Molecular Mus Muscle Cells Nature Oligonucleotides Organogenesis Pathogenesis Pathology Polymerase Proteins RNA Polymerase II Regulation Role Sequence Analysis Spliced Genes Technology Therapeutic Time Transcription Factor TFIIB Transcriptional Regulation Up-Regulation Work chromatin immunoprecipitation deep sequencing genome-wide locked nucleic acid overexpression pressure promoter skeletal tool transcription factor

项目摘要

DESCRIPTION (provided by applicant): Cardiac hypertrophy is characterized by a generalized increase in gene expression that is commensurate with the increase in myocyte size and mass, on which is superimposed more robust changes in the expression of specialized genes. While transcriptional regulation of some of those genes has been validated, we do not have comprehensive, genome-wide, knowledge of which genes are regulated by transcription vs. those that may be independently regulated by posttranscriptional mechanisms involving microRNA. Additionally, we do not know the mode of transcriptional regulation - de novo RNA polymerase II (pol II) recruitment vs. the release of paused pol II - or the regulators involved. One of the earliest changes observed after applying pressure overload on a mouse heart, is the downregulation of miR-1, which precedes any other miRNA changes, the increase in cardiac mass, or contractile dysfunction. This suggested that miR-1 might be a cause rather than an effect of the underlying pathogenesis. Our preliminary data show that miR-1 targets two major components of basic transcription, general transcription factor 2B (TFIIB) and cyclin-dependent kinase 9 (Cdk9), which are the key regulators of pol II recruitment and elongation, respectively. Inhibiting miR-1 with locked nucleic acid- modified anti-miR oligo in the heart is sufficient for inducing upregulation of these targets, and conversely overexpression of miR-1 suppresses their expression. More significantly, chromatin immunoprecipitation-deep sequencing analysis (ChIP-Seq) reveals that supplementing cardiac myocytes with miR-1 suppresses de novo pol II recruitment on a subset of genes, while inducing pausing on another, in concordance with its suppression of TFIIB and Cdk9, respectively. The same subsets of genes are inversely regulated during cardiac hypertrophy as miR-1 is downregulated. In general, the regulation by de novo pol II recruitment and that by the release of promoter-paused pol II seem to be mutually exclusive. The former mainly regulates ~6% of genes mainly including those with specialized functions (e.g. contractile, extracellular matrix, immune response...etc), while the latter involve ~ 25% of expressed genes mainly including housekeeping/essential genes (e.g. protein and mRNA turnover genes, basal transcription factor, splicing genes...etc). These results led to the hypotheses for this grant. i- Downregulation of miR-1 is required for upregulation of TFIIB and Cdk9 during cardiac hypertrophy and, accordingly, the associated changes in gene expression. ii- Selective inhibition of TFIIB in the heart during cardiac hypertrophy will inhibit de novo recruitment of pol II to the promoters of a subset of genes (~6%) including those involved in the development of cardiomyopathy (e.g. ANF, BNP, alpha skeletal actin, collagen, etc.) This will not inhibit the increase in cardiac mass but will ameliorate contractile dysfunction during hypertrophy. iii- Selective inhibition of Cdk9 in the heart during cardiac hypertrophy will inhibit promoter clearance of paused pol II on all essential/housekeeping genes (~25%) in the heart (e.g. Vdac1, pinin, TFIIB, Cdk9, MAPK1, etc). This will inhibit the increase in cardiac mass and result in precipitous cardiac failure. The specific aims are 1) Identify the mechanisms involved in the regulation of TFIIB and Cdk9, and basic gene transcription, during cardiac hypertrophy. 2) Determine the role of TFIIB in gene transcription and the development of cardiac hypertrophy. 3) Determine the role of Cdk9 in gene transcription and the development of cardiac hypertrophy.

描述（由申请人提供）：心脏肥大的特征是基因表达普遍增加，与肌细胞大小和质量的增加相称，其上叠加了特殊基因表达的更强烈变化。虽然其中一些基因的转录调控已经得到验证，但我们并没有全面的、全基因组范围的知识，了解哪些基因受转录调控，哪些基因可能受涉及 microRNA 的转录后机制独立调控。此外，我们不知道转录调控的模式 - 从头 RNA 聚合酶 II (pol II) 招募与暂停的 pol II 的释放 - 或所涉及的调节因子。对小鼠心脏施加压力超负荷后观察到的最早变化之一是 miR-1 的下调，它先于任何其他 miRNA 的变化、心脏质量的增加或收缩功能障碍。这表明 miR-1 可能是潜在发病机制的原因而不是结果。我们的初步数据表明，miR-1 靶向基本转录的两个主要成分，即通用转录因子 2B (TFIIB) 和细胞周期蛋白依赖性激酶 9 (Cdk9)，它们分别是 pol II 募集和延伸的关键调节因子。在心脏中用锁核酸修饰的抗miR寡核苷酸抑制miR-1足以诱导这些靶标的上调，并且相反，miR-1的过表达抑制它们的表达。更重要的是，染色质免疫沉淀-深度测序分析 (ChIP-Seq) 表明，用 miR-1 补充心肌细胞可抑制一部分基因上的 pol II 从头招募，同时诱导另一部分基因暂停，分别与其对 TFIIB 和 Cdk9 的抑制一致。当 miR-1 下调时，相同的基因子集在心脏肥大期间受到反向调节。一般来说，从头招募 pol II 的调节和启动子暂停的 pol II 释放的调节似乎是相互排斥的。前者主要调控约6%的基因，主要包括具有特殊功能的基因（例如收缩、细胞外基质、免疫反应等），而后者涉及约25%的表达基因，主要包括管家/必需基因（例如蛋白质和mRNA转换基因、基础转录因子、剪接基因等）。这些结果引出了本次资助的假设。 i- 心脏肥大期间 miR-1 的下调是 TFIIB 和 Cdk9 上调所必需的，因此，基因表达的相关变化也是如此。 ii- 在心脏肥大期间选择性抑制心脏中的 TFIIB 将抑制 pol II 从头招募到一部分基因 (~6%) 的启动子，包括那些参与心肌病发展的基因（例如 ANF、BNP、α 骨骼肌动蛋白、胶原蛋白等）。这不会抑制心脏质量的增加，但会改善心脏肥大期间的收缩功能障碍。肥大。 iii- 心脏肥大期间选择性抑制心脏中的 Cdk9 将抑制启动子清除心脏中所有必需/管家基因 (~25%) 上暂停的 pol II（例如 Vdac1、pinin、TFIB、Cdk9、MAPK1 等）。这将抑制心脏质量的增加并导致急剧的心力衰竭。具体目标是 1) 确定涉及的机制心脏肥大期间 TFIIB 和 Cdk9 的调节以及基本基因转录。 2)确定TFIIB在基因转录和心脏肥大发展中的作用。 3)确定Cdk9在基因转录和心脏肥大发展中的作用。