Microglia, resident macrophages of the brain that act as primary immune cells, play essential roles in innate immunity and neuroinflammatory pathologies. Microglial cells are rapidly activated in response to infection and inflammation/injury, associated with the expression of proinflammatory genes and secretion of cytokines. The bromodomain and extra-terminal (BET) inhibitor JQ1 has been shown to be an epigenetic agent that reduces inflammation. In this study, we investigated the mechanisms underlying the anti-inflammatory and anti-migratory functions of JQ1 and the genes targeted by JQ1 in lipopolysaccharide (LPS)-activated human microglial clone 3 (HMC3) cells using RNA-sequencing (RNA-seq). We analyzed the pattern of inflammation-related genes (chemokines, cytokines, and interferon-stimulated genes) and migration-related genes with JQ1 treatment from differentially expressed genes analysis in HMC3 cells. We found that LPS-induced IRF1 directly regulated inflammation- and migration-related genes and that JQ1 significantly reduced IRF1 and its target genes. Additionally, IRF1 attenuation significantly downregulated target genes and inhibited microglial migration. Our data suggest that the BET inhibitor JQ1 can modulate the inflammatory response and migration through the regulation of LPS-induced IRF1 in human microglia.

TRIM71 (tripartite motif-containing 71) belongs to the TRIM-NHL protein family, which plays a conserved role in regulating early development and differentiation. However, the molecular functions of TRIM71 have remained largely unknown. Here, we explored the role of TRIM71 together with modulation of Lin28B-let-7-HMGA2 (high-mobility group AT-hook 2) signaling in tumorigenesis. TRIM71 overexpression opposed Lin28B-induced transformation in primary cells and inhibited tumor formation in a mouse model. Specific knockdown of TRIM71 expression increased cancer cell proliferation and invasion. Conversely, overexpression of wild-type TRIM71 in non-small cell lung carcinoma (NSCLC) cells in which Lin28B-let-7-HMGA2 signaling was conserved decreased both cancer cell phenotypes. More importantly, overexpression of an ubiquitin transfer activity-deficient TRIM71 mutant in NSCLC cells had no effect on proliferation or invasion, regardless of the conservation status of Lin28B-let-7-HMGA2 signaling. The tumorigenic inhibitory action of TRIM71 was antagonized by overexpression of the TRIM71 downstream targets, Lin28B and HMGA2. Furthermore, a bioinformatics analysis revealed that TRIM71 expression was downregulated in various types of cancer tissue from patients. Taken together, these data indicate that TRIM71 acts through post-transcriptional repression of Lin28B and subsequent modulation of let-7-HMGA2 signaling during tumorigenesis to potentially function as a tumor suppressor.

Hepatocellular carcinoma (HCC) is one of the most prevalent and poorly responsive cancers worldwide. Bromodomain and extraterminal (BET) inhibitors, such as JQ1 and OTX-015, inhibit BET protein binding to acetylated residues in histones. However, the physiological mechanisms and regulatory processes of BET inhibition in HCC remain unclear. To explore BET inhibitors’ potential role in the molecular mechanisms underlying their anticancer effects in HCC, we analyzed BET inhibitor-treated HCC cells’ gene expression profiles with RNA-seq and bioinformatics analysis. BET inhibitor treatment significantly downregulated genes related to bromodomain-containing proteins 4 (BRD4), such as ACSL5, SLC38A5, and ICAM2. Importantly, some cell migration-related genes, including AOC3, CCR6, SSTR5, and SCL7A11, were significantly downregulated. Additionally, bioinformatics analysis using Ingenuity Knowledge Base Ingenuity Pathway Analysis (IPA) revealed that SMARCA4 regulated migration response molecules. Furthermore, knockdown of SMARCA4 gene expression by siRNA treatment significantly reduced cell migration and the expression of migration-related genes. In summary, our results indicated that BET inhibitor treatment in HCC cell lines reduces cell migration through the downregulation of SMARCA4.

Background: Toll-like receptor 4 (TLR4) ligands such as lipopolysaccharide (LPS) activate immunomodulatory functions and the migration of human mesenchymal stromal cells (hMSCs). Here, we study the migration-related gene expression of LPS-stimulated hMSCs and the role and regulation of one of the upregulated genes, encoding the interferon-induced transmembrane protein 1 (IFITM1).Methods: Gene expression profiles were determined by whole-transcriptome analysis (RNA-seq) and quantitative real-time PCR (qRT-PCR). Bioinformatics approaches were used to perform network and pathway analyses. The cell migration-related genes were identified with an in vitro wound healing assay. RNA interference (RNAi) was used to suppress the IFITM1 gene expression. The IFITM1 gene enhancer was analyzed by chromatin immunoprecipitation (ChIP) sequencing, ChIP-to-PCR, luciferase reporter assays, and qRT-PCR for enhancer RNAs (eRNAs).Results: RNA-seq confirmed IFITM1 as an LPS-stimulated gene, and RNAi demonstrated its importance for the LPSstimulated migration. LPS treatment increased the eRNA expression in enhancer region R2 (2 kb upstream) of the IFITM1 gene and enriched R2 for H3K27ac. Bioinformatics implicated the transcription factors NF-κB and IRF1, ChIP assays revealed their binding to R2, and chemical inhibition of NF-κB and RNAi directed against IRF1 prevented R2 eRNA and IFITM1 gene expression. Conclusions: Increased expression of the IFITM1 gene is required for LPS-stimulated hMSC migration. We described several underlying changes in the IFITM1 gene enhancer, most notably the NF-κB-mediated activation of enhancer region R2.

Hepatocellular carcinoma (HCC) is the most common primary liver cancer and poor prognosis. Emerging evidence suggests that epigenetic alterations play a crucial role in HCC, suggesting epigenetic inhibition as a promising therapeutic approach. Indeed, the bromodomain and extra-terminal (BET) inhibitors inhibit the proliferation and invasion of various cancers but still lack a strong mechanistic rationale. Here, we identified the differentially expressed mRNAs (DEmRNAs) and lncRNAs (DElncRNAs) in human HCC cell line HepG2 treated with the BET inhibitors, JQ1, OTX015, or ABBV-075. We analyzed the correlation between DEmRNAs and DElncRNAs in common for the three inhibitors based on their expression profiles and performed functional annotation pathway enrichment analysis. Most of these shared DEmRNAs and DElncRNAs, including some novel transcripts, were downregulated, indicating decreased proliferation/adhesion and increased apoptosis/inflammation. Our study suggests that BET proteins play a crucial role in regulating cancer progression-related genes and provide a valuable resource for novel putative biomarkers and therapeutic targets in HCC.