Regulation of Transcription by Poly(Adp-Ribose) Polymerase-1 in Adipogenesis and Inflammation

Date

2013-11-20

Authors

Luo, Xin

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Abstract

Poly(ADP-ribose) polymerases (PARPs) comprise a family of enzymes sharing a conserved catalytic domain that supports mono- or poly(ADP-ribosyl)transferase activity. PARP-1 is the founding member and controls transcriptional outcomes by modulating chromatin structure and the activity of transcriptional regulators. PARP-1 plays a role in various biological processes including differentiation, metabolism, inflammation and hormonal response and is associated with many pathological conditions including cancer, infection, cardiovascular disorders, and metabolic deregulation. However, the molecular mechanisms of how PARP-1 regulates transcription in different systems are unknown. In my thesis, I have investigated the transcriptional regulation by PARP-1 in both normal differentiation program and inflammatory stress response. To determine the role of PARP-1 in adipogenesis, I used a variety of cell-based and molecular assays in 3T3-L1 preadipocytes. I found that the nuclear levels of PARylation fluctuate during adipogenesis and that PARP-1 represses adipogenesis by modulating the transcription of key adipogenic factors, C/EBPα and PPARγ2. I further showed that PARP-1 represses the activity of the early transcription factor C/EBPβ through Poly(ADP-ribosyl)ation (PARylation) at specific sites. Collectively, my study explored PARP-1’s role as a transcriptional repressor through modulating C/EBPβ, during adipogenesis. In order to study how inflammation affects cardiac function and whether PARP-1 plays a role in this process, I characterized the transcriptome of an immortalized adult human ventricular cardiomyocyte cell line (AC16) in response to tumor necrosis factor (TNFα) stimulation. Using genomic approaches including global nuclear run-on sequencing (GRO-seq) and chromatin immunoprecipitation coupled with sequencing (ChIP-seq), I identified ~30,000 transcribed regions including protein coding and non coding transcripts generated from RNA polymerase I, II and III. In addition, I observed that AC16 cells rapidly and dynamically reorganize their transcriptomes in response to TNFα stimulation in an NF-κB-dependent manner, switching from a basal state to a proinflammatory state, affecting a spectrum of cardiac-associated protein-coding and non-coding genes. I observed distinct Pol II dynamics for up- and downregulated genes. Furthermore, I found that PARP-1 is enriched at target genes corresponding to gene expression level, and facilitates NF-κB regulatory circuits. Overall, my study helped to clarify the connection between inflammation and cardiomyocyte function at the transcriptional level.

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