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Dynamic reorganization of the AC16 cardiomyocyte transcriptome in response to TNF? signaling revealed by integrated genomic analyses.

ABSTRACT: Defining cell type-specific transcriptomes in mammals can be challenging, especially for unannotated regions of the genome. We have developed an analytical pipeline called groHMM for annotating primary transcripts using global nuclear run-on sequencing (GRO-seq) data. Herein, we use this pipeline to characterize the transcriptome of an immortalized adult human ventricular cardiomyocyte cell line (AC16) in response to signaling by tumor necrosis factor alpha (TNF?), which is controlled in part by NF-?B, a key transcriptional regulator of inflammation. A unique aspect of this work is the use of the RNA polymerase II (Pol II) inhibitor ?-amanitin, which we used to define a set of RNA polymerase I and III (Pol I and Pol III) transcripts.Using groHMM, we identified ~30,000 coding and non-coding transcribed regions in AC16 cells, which includes a set of unique Pol I and Pol III primary transcripts. Many of these transcripts have not been annotated previously, including enhancer RNAs originating from NF-?B binding sites. In addition, we observed that AC16 cells rapidly and dynamically reorganize their transcriptomes in response to TNF? stimulation in an NF-?B-dependent manner, switching from a basal state to a proinflammatory state affecting a spectrum of cardiac-associated protein-coding and non-coding genes. Moreover, we observed distinct Pol II dynamics for up- and downregulated genes, with a rapid release of Pol II into productive elongation for TNF?-stimulated genes. As expected, the TNF?-induced changes in the AC16 transcriptome resulted in corresponding changes in cognate mRNA and protein levels in a similar manner, but with delayed kinetics.Our studies illustrate how computational genomics can be used to characterize the signal-regulated transcriptome in biologically relevant cell types, providing new information about how the human genome is organized, transcribed and regulated. In addition, they show how ?-amanitin can be used to reveal the Pol I and Pol III transcriptome. Furthermore, they shed new light on the regulation of the cardiomyocyte transcriptome in response to a proinflammatory signal and help to clarify the link between inflammation and cardiomyocyte function at the transcriptional level.

SUBMITTER: Luo X

PROVIDER: S-EPMC3945043 | biostudies-literature | 2014 Feb

REPOSITORIES: biostudies-literature

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Dynamic reorganization of the AC16 cardiomyocyte transcriptome in response to TNFα signaling revealed by integrated genomic analyses.

Luo Xin X Chae Minho M Krishnakumar Raga R Danko Charles G CG Kraus W Lee WL

BMC genomics 20140224

<h4>Background</h4>Defining cell type-specific transcriptomes in mammals can be challenging, especially for unannotated regions of the genome. We have developed an analytical pipeline called groHMM for annotating primary transcripts using global nuclear run-on sequencing (GRO-seq) data. Herein, we use this pipeline to characterize the transcriptome of an immortalized adult human ventricular cardiomyocyte cell line (AC16) in response to signaling by tumor necrosis factor alpha (TNFα), which is co ...[more]

PMID: 24564208

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Project description:BackgroundRadix Ardisia (Jab Bik Lik Jib) is a common Miao medicine and is widely distributed in the Guizhou region of southern China. The botanical origin of Radix Ardisia includes the dry root and rhizome of Ardisia Crenata Sims (ACS) or Ardisia Crispa (Thunb.) A.DC. (AC), which are closely related species morphologically. However, the secondary metabolites in their roots are different from one another, especially the flavonoids, and these differences have not been thoroughly explored at the molecular level. This project preliminarily identified regulatory molecular mechanisms in the biosynthetic pathways of the flavonoids between ACS and AC using a multi-omics association analysis.MethodsIn this study, we determined the total levels of saponin, flavonoid, and phenolic in Radix Ardisia from different origins. Integrated transcriptome and metabolome analyses were used to identify the differentially expressed genes (DEGs) and differentially expressed metabolites (DEM). We also performed conjoint analyses on DEGs and DEMs to ascertain the degree pathways, and explore the regulation of flavonoid biosynthesis.ResultsThe total flavonoid and phenolic levels in ACS were significantly higher than in AC (P < 0.05). There were 17,685 DEGs between ACS vs. AC, 8,854 were upregulated and 8,831 were downregulated. Based on this, we continued to study the gene changes in the flavonoid biosynthesis pathway, and 100 DEGs involving flavonoid biosynthesis were differentially expressed in ACS and AC. We validated the accuracy of the RNA-seq data using qRT-PCR. Metabolomic analyses showed that 11 metabolites were involved in flavonoid biosynthesis including: Naringenin, Luteolin, Catechin, and Quercetin. A conjoint analysis of the genome-wide connection network revealed the differences in the types and levels of flavonoid compounds between ACS and AC. The correlation analysis showed that Naringenin, Luteolin, Catechin, and Quercetin were more likely to be key compounds in the flavonoid biosynthesis pathway also including 4CL, AOMT, CHS, CHI, DFR, F3'5'H, FLS, and LAR.ConclusionsThis study provides useful information for revealing the regulation of flavonoid biosynthesis and the regulatory relationship between metabolites and genes in the flavonoid biosynthesis pathway in Radix Ardisia from different origins.

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Dynamic reorganization of the AC16 cardiomyocyte transcriptome in response to TNF? signaling revealed by integrated genomic analyses.

Publications

Dynamic reorganization of the AC16 cardiomyocyte transcriptome in response to TNFα signaling revealed by integrated genomic analyses.

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