Mdivi-1: Effective but complex mitochondrial fission inhibitor.

Mdivi-1, Mitochondrial DIVIsion inhibitor 1, has been widely employed in research under the assumption that it exclusively influences mitochondrial fusion, but effects other than mitochondrial dynamics have been underinvestigated. This paper provides transcriptome and DNA methylome-wide analysis for Mdivi-1 treated SH-SY5Y human neuroblastoma cells using RNA sequencing (RNA-seq) and methyl capture sequencing (MC-seq) methods. Gene ontology analysis of RNA sequences revealed that p53 transcriptional gene network and DNA replication initiation-related genes were significantly up and down-regulated, respectively, showing the correlation with the arrest cell cycle in the G1 phase. MC-seq, a powerful sequencing method for capturing DNA methylation status in CpG sites, revealed that although Mdivi-1 does not induce dramatic DNA methylation change, the subtle alterations were concentrated within the CpG island. Integrative analysis of both sequencing data disclosed that the p53 transcriptional network was activated while the Parkinson's disease pathway was halted. Next, we investigated several changes in mitochondria in response to Mdivi-1. Copy number and transcription of mitochondrial DNA were suppressed. ROS levels increased, and elevated ROS triggered mitochondrial retrograde signaling rather than inducing direct DNA damage. In this study, we could better understand the molecular network of Mdivi-1 by analyzing DNA methylation and mRNA transcription in the nucleus and further investigating various changes in mitochondria, providing inspiration for studying nuclear-mitochondrial communications.

Epigenetic landscape analysis reveals the significance of early reduced chromatin accessibility in osteoclastogenesis.

Recently improved techniques could provide snapshots of chromatin structure generated based on chromatin accessibility. Since chromatin accessibility determines transcriptional potential, it has been attempted in a variety of cell systems. However, there has been no genome-wide analysis of chromatin accessibility for the entire murine osteoclast (OC) differentiation process. We performed an Assay for Transposase-Accessible Chromatin (ATAC)-sequencing (seq) during RANKL-induced OC differentiation and found that global chromatin accessibility decreased, especially early in OC differentiation. The global histone H3K27Ac level, an active histone modification mark, was diminished during OC differentiation by western blot and histone extract experiments. Its genomic enrichment was also reduced based on publicly available H3K27Ac chromatin immunoprecipitation (ChIP)-seq data. ATAC-seq and H3K27Ac ChIP-seq data demonstrated that RANKL induced a less accessible chromatin state during OC differentiation. Restoration of reduced H3K27Ac, presumably representing accessible states upon acetate treatment, suppresses OC differentiation by provoking immune-related gene expression. Subsequential integrative analysis of ATAC-seq, RNA-seq after acetate treatment, and H3K27Ac ChIP-seq reveals that Irf8 and its downstream targets are the most vulnerable to chromatin accessibility changes and acetate supplementation. Taken together, our study generated chromatin accessibility maps during the whole OC differentiation and suggested perturbation of chromatin accessibility might be a potential therapeutic strategy for excessive OC diseases.

Nucleoporin 210 Serves a Key Scaffold for SMARCB1 in Liver Cancer.

The roles of chromatin remodelers and their underlying mechanisms of action in cancer remain unclear. In this study, SMARCB1, known initially as a bona fide tumor suppressor gene, was investigated in liver cancer. SMARCB1 was highly upregulated in patients with liver cancer and was associated with poor prognosis. Loss- and gain-of-function studies in liver cells revealed that SMARCB1 loss led to reduced cell proliferation, wound healing capacity, and tumor growth in vivo. Although upregulated SMARCB1 appeared to contribute to switch/sucrose nonfermentable (SWI/SNF) complex stability and integrity, it did not act using its known pathways antagonism with EZH2 or association between TP53 or AMPK. SMARCB1 knockdown induced a mild reduction in global H3K27 acetylation, and chromatin immunoprecipitation sequencing of SMARCB1 and acetylated histone H3K27 antibodies before and after SMARCB1 loss identified Nucleoporin210 (NUP210) as a critical target of SMARCB1, which bound its enhancer and changed H3K27Ac enrichment and downstream gene expression, particularly cholesterol homeostasis and xenobiotic metabolism. Notably, NUP210 was not only a putative tumor supporter involved in liver cancer but also acted as a key scaffold for SMARCB1 and P300 to chromatin. Furthermore, SMARCB1 deficiency conferred sensitivity to doxorubicin and P300 inhibitor in liver cancer cells. These findings provide insights into mechanisms underlying dysregulation of chromatin remodelers and show novel associations between nucleoporins and chromatin remodelers in cancer. SIGNIFICANCE: This study reveals a novel protumorigenic role for SMARCB1 and describes valuable links between nucleoporins and chromatin remodelers in cancer by identifying NUP210 as a critical coregulator of SMARCB1 chromatin remodeling activity.

SMARCB1 Acts as a Quiescent Gatekeeper for Cell Cycle and Immune Response in Human Cells.

Switch/sucrose non-fermentable (SWI/SNF)-related matrix-associated actin-dependent regulator of chromatin (SMARC) subfamily B member 1 (SMARCB1) is a core subunit of the switch/sucrose non-fermentable (SWI/SNF) complex, one of the adenosine triphosphate (ATP)-dependent chromatin remodeler complexes. The unique role of SMARCB1 has been reported in various cellular contexts. Here, we focused on the general role of the ubiquitous expression of SMARCB1 in a normal cell state. We selected ARPE19 (human primary retinal pigment epithelium) and IMR90 (from human fetal lung fibroblasts) cell lines as they have completely different contexts. Furthermore, although these cell lines have been immortalized, they are relatively close to normal human cells. The loss of SMARCB1 in ARPE19 and IMR90 cells reduced cell cycle progression via the upregulation of P21. Transcriptome analysis followed by SMARCB1 knockdown in both cell lines revealed that SMARCB1 was not only involved in cell maintenance but also conferred immunomodulation. Of note, SMARCB1 bound to interleukin (IL) 6 promoter in a steady state and dissociated in an active immune response state, suggesting that SMARCB1 was a direct repressor of IL6, which was further confirmed via loss- and gain-of-function studies. Taken together, we demonstrated that SMARCB1 is a critical gatekeeper molecule of the cell cycle and immune response.

Oncogenic IL7R is downregulated by histone deacetylase inhibitor in esophageal squamous cell carcinoma via modulation of acetylated FOXO1.

The interleukin-7 receptor (IL7R) is generally expressed in immune cells and is critical in survival, development and homeostasis in the immune system. Advanced genome-wide cancer studies have reported that IL7R is genetically amplified in human esophageal squamous cell carcinoma (ESCC), however, the exact role of IL7R in ESCC has not been investigated. In the present study, it was found that IL7R was overexpressed in ESCC cohorts and the loss of IL7R induced anti-oncogenic effects in ESCC cell lines. A small panel of epigenetic drugs were screened for their ability to downregulate the expression of IL7R. Unexpectedly, apicidin, a histone deacetylase (HDAC) inhibitor, effectively downregulated the expression of IL7R in a dose-dependent manner at an early time-point, as determined by quantitative polymerase chain reaction and IL7R immunostaining, and did not require de novo protein synthesis. Of note, apicidin induced the acetylation of Forkhead box-containing protein, O subfamily 1, which acts as a repressor at the IL7R promoter, accompanied with depleted active histone modifications based on chromatin immunoprecipitation assay. Taken together, these results demonstrated that targeting oncogenic IL7R in ESCC by HDAC inhibitors may be a valuable therapeutic approach.

Epigenetic landscape change analysis during human EMT sheds light on a key EMT mediator TRIM29.

Epithelial to mesenchymal transition (EMT) is a key trans-differentiation process, which plays a critical role in physiology and pathology. Although gene expression changes in EMT have been scrutinized, study of epigenome is in its infancy. To understand epigenetic changes during TWIST-driven EMT, we used the AcceSssIble assay to study DNA methylation and chromatin accessibility in human mammary epithelial cells (HMECs). The DNA methylation changes were found to have functional significance in EMT - i.e. methylated genes were enriched for E-box motifs that can be recognized by TWIST, at the promoters suggesting a potential targeting phenomenon, whereas the demethylated regions were enriched for pro-metastatic genes, supporting the role of EMT in metastasis. TWIST-induced EMT triggers alterations in chromatin accessibility both independent of and dependent on DNA methylation changes, primarily resulting in closed chromatin conformation. By overlapping the genes, whose chromatin structure is changed during early EMT and a known "core EMT signature", we identified 18 driver candidate genes during EMT, 14 upregulated and 4 downregulated genes with corresponding chromatin structure changes. Among 18 genes, we focused on TRIM29 as a novel marker of EMT. Although loss of TRIM29 is insufficient to suppress CDH, it is enough to induce CDH2 and VIM. Gene functional annotation analysis shows the involvement of TRIM29 in epidermal development, cell differentiation and cell migration. Taken together, our results provide a robust snapshot of chromatin state during human EMT and identify TRIM29 as a core mediator of EMT.

Epigenetic reader BRD4 inhibition as a therapeutic strategy to suppress E2F2-cell cycle regulation circuit in liver cancer.

Deregulation of the epigenome component affects multiple pathways in the cancer phenotype since the epigenome acts at the pinnacle of the hierarchy of gene expression. Pioneering work over the past decades has highlighted that targeting enzymes or proteins involved in the epigenetic regulation is a valuable approach to cancer therapy. Very recent results demonstrated that inhibiting the epigenetic reader BRD4 has notable efficacy in diverse cancer types. We investigated the potential of BRD4 as a therapeutic target in liver malignancy. BRD4 was overexpressed in three different large cohort of hepatocellular carcinoma (HCC) patients as well as in liver cancer cell lines. BRD4 inhibition by JQ1 induced anti-tumorigenic effects including cell cycle arrest, cellular senescence, reduced wound healing capacity and soft agar colony formation in liver cancer cell lines. Notably, BRD4 inhibition caused MYC-independent large-scale gene expression changes in liver cancer cells. Serial gene expression analyses with SK-Hep1 liver cancer cells treated with JQ1 to delineate the key player of BRD4 inhibition identified E2F2 as the first line of downstream direct target of BRD4. Further experiments including chromatin immunoprecipitation (ChIP) assay and loss of function study confirmed E2F2 as key player of BRD4 inhibition. Overexpressed E2F2 is a crucial center of cell cycle regulation and high expression of E2F2 is significantly associated with poor prognosis of HCC patients. Our findings reveal BRD4-E2F2-cell cycle regulation as a novel molecular circuit in liver cancer and provide a therapeutic strategy and innovative insights for liver cancer therapies.

JQ1, an inhibitor of the epigenetic reader BRD4, suppresses the bidirectional MYC-AP4 axis via multiple mechanisms.

Bromodomain and extra-terminal domain (BET) family proteins are representative epigenetic modulators that read acetylated lysine residues and transfer cellular signals. Recently, the BET protein inhibitor JQ1 was developed and has been extensively studied in many cancer cell types. We demonstrated that JQ1 effectively suppressed the MYC-AP4 axis and induced antitumorigenic effects by targeting a bidirectional positive loop between MYC and AP4 which was first proposed in the present study. MYC and AP4 are the direct targets of BRD4, as demonstrated by chromatin immunoprecipitation (ChIP) assay and BRD4 loss-of-function experiments. Although inhibition of the MYC/MAC dimer suppressed AP4, the efficacy of suppression was not as effective as BRD4 inhibition. Notably, AP4 loss-of-function studies demonstrated that AP4 is a major critical target of JQ1 and that MYC is a novel downstream target of AP4, as demonstrated by AP4 binding to the MYC promoter. Taken together, our results suggest that the epigenetic reader BRD4 is a key mediator of the activated MYC-AP4 axis, which supports the possibility that targeting BET protein is a novel therapeutic strategy for MYC-AP4 axis-activated cancers.

Myotonic dystrophy type I combined with X-linked dominant Charcot-Marie-Tooth neuropathy.

Both the myotonic dystrophy type 1 (DM1) and the X-linked dominant Charcot-Marie-Tooth disease (CMTX1) are well-established inherited neuromuscular disorders characterized by progressive weakness and atrophy of the distal limb muscles. The underlying causes of the DM1 and CMTX1 are mutations in the DMPK and GJB1 gene, respectively. A patient with both DM1 and CMTX1 inherited these from his father and mother, respectively. Histopathological and electrodiagnostic studies revealed both chronic neuropathic and myopathic features. Physical disabilities were more severe than seen with either DM1 or CMTX1 alone. In addition, the present case reveals an asymmetric atrophy (22%) of the right calf muscle compared to the left side.