Epigenetic regulation of gene expression by BRD4 in cancer and innate immune response

Dong, Xingchen

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https://hdl.handle.net/2142/108583 Copy

Description

Title

Epigenetic regulation of gene expression by BRD4 in cancer and innate immune response

Author(s)

Dong, Xingchen

Issue Date

2020-07-14

Director of Research (if dissertation) or Advisor (if thesis)

Chen, Lin-feng

Doctoral Committee Chair(s)

Chen, Lin-feng

Committee Member(s)

• Chen, Jie
• Huang, Raven
• Kalsotra, Auinash

Department of Study

Biochemistry

Discipline

Biochemistry

Degree Granting Institution

University of Illinois at Urbana-Champaign

Degree Name

Ph.D.

Degree Level

Dissertation

Keyword(s)

• epigenetic
• BET inhibition
• BRD4
• miRNA
• cancer
• innate immune response
• Salmonella
• NLRC4 inflammasome
• transcriptional regulation

Abstract

Epigenetic regulation of gene expression plays essential roles in controlling normal cellular functions as well as abnormal cellular activities in human diseases like inflammatory diseases and cancers. The bromodomain-containing protein 4 (BRD4), one of the BET (bromodomain and extra terminal) family proteins, represents a class of epigenetic readers that regulate gene transcription by binding to acetylated lysine of histone and non-histone proteins via their bromodomains (BDs). This interaction either changes the availabilities of cis-regulatory elements by altering chromatin compacity or activates specific transcription factors, both of which contribute to transcriptional activation of BRD4’s target genes. BRD4 has been implicated in the dysregulated transcription of oncogenes and inflammatory genes during disease progression; thus, the small molecule inhibitors that target the bromodomains of BRD4, as well as other BET family proteins, are under clinical investigations. Gastric cancer has become one of the leading malignancies that cause death with a 5-year survival rate of 10%. Although BET inhibitors (BETis) have shown potential therapeutic effects against gastric cancer, the detailed mechanism by which BRD4 facilities gastric cancer cell proliferation remains elusive. In an effort to understand the contribution of BET to gastric cancer development, we confirmed that BRD4 was overexpressed in gastric cancer patient tissues compared to normal tissue. Meanwhile, BET inhibitor JQ1 inhibited multiple gastric cancer cell proliferation by inducing BRD4-dependent cellular senescence. Depletion of BRD4, but not other BET proteins, recapitulated JQ1-induced cellular senescence with increased cellular SA--Galactose activity and elevated p21 levels. BRD4 inhibited p21 expression at the post-transcriptional level through activating the transcription of miR-106b-5p, which targets the 3’-UTR of p21 mRNA. Overexpression of miR-106b-5p prevented JQ1-induced p21 expression and BRD4 inhibition-associated cellular senescence, whereas miR-106b-5p inhibitor upregulated p21 and induced cellular senescence. Finally, we demonstrated that inhibition of E2F suppressed the binding of BRD4 to the promoter of miR-106b-5p and inhibited its transcription, leading to the increased p21 levels and cellular senescence in gastric cancer cells. Our results reveal a novel mechanism by which BRD4 regulates cancer cell proliferation by modulating the cellular senescence through the E2F/miR-106b-5p/p21 axis and provide new insights into using BET inhibitors as potential anti-cancer drugs. Other than its critical role in facilitating tumor cell proliferation, BRD4 has also been investigated in the dysregulated cytokine production at the transcriptional level during inflammatory disease progression. BRD4 has been shown to activate NF-B-dependent inflammatory gene expression by binding to acetylated RelA subunit of NF-B through its bromodomains, highlighting its essential role in transcriptional regulation of cytokine production. Although the production of inflammatory cytokines is heavily regulated at the transcriptional level, a group of functional cytokines, such as interleukin-1 (IL-1) and IL-18, require post-translational maturation mediated by the activation of the inflammasome complex. In order to study the comprehensive role of BRD4 in cytokine production, we seek to investigate if BRD4 plays a role in regulating inflammasome activation and cytokine maturation. Using Salmonella enterica serovar Typhimurium (S. Typhimurium) as a NLRC4 inflammasome activator, we found that Brd4-deficient bone marrow-derived macrophages (BMDMs) displayed blunted NLRC4 inflammasome activation with decreased caspase-1 activation, Asc oligomerization, IL-1 maturation, gasdermin-D (Gsdmd) cleavage, and pyroptosis upon bacterial infection. RNA-seq results unveiled that the ablation of Brd4 in BMDMs suppressed the transcription of Naips and Nlrc4. Mechanistically, ChIP-seq analysis revealed that Brd4 co-localized with Irf8 and Pu.1 at the promoter regions of Naips, activating their transcriptions. Moreover, myeloid lineage-specific Brd4 conditional-knockout (Brd4-CKO) mice were more sensitive to S. Typhimurium infection with the significantly enhanced bacterial burden and tissue damages. Altogether, our findings emphasize the unexpected role of Brd4 in NLRC4 inflammasome activation through modulating the transcription of Naips and Nlrc4. ¬¬ In summary, BRD4, as an epigenetic reader, activates the transcription of diverse sets of genes through engaging distinct transcription factors in the cell context- and disease-dependent manner. In the gastric cancer cells, BRD4 controls cancer cell proliferation by modulating the cellular senescence through the E2F/miR-106b-5p/p21 axis. Meanwhile, in the innate immune response, Brd4 engages Irf8 and Pu.1 in maintaining the transcription of Naips and Nlrc4, which confers a prompt inflammasome activation against invading pathogens. Of note, mechanistically, BRD4’s transcriptional activities highly, but not exclusively, depend on the interaction between its bromodomains and acetylated lysine on histone or transcription factors, which supports the clinical investigations using BETis as anti-cancer and -inflammatory disease drugs.

Graduation Semester

2020-08

Type of Resource

Thesis

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Copyright 2020 Xingchen Dong

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