The SWI/SNF Subunit/Tumor Suppressor BAF47/INI1 Is Essential in Cell Cycle Arrest upon Skeletal Muscle Terminal Differentiation

Myogenic terminal differentiation is a well-orchestrated process starting with permanent cell cycle exit followed by muscle-specific genetic program activation. Individual SWI/SNF components have been involved in muscle differentiation. Here, we show that the master myogenic differentiation factor MyoD interacts with more than one SWI/SNF subunit, including the catalytic subunit BRG1, BAF53a and the tumor suppressor BAF47/INI1. Downregulation of each of these SWI/SNF subunits inhibits skeletal muscle terminal differentiation but, interestingly, at different differentiation steps and extents. BAF53a downregulation inhibits myotube formation but not the expression of early muscle-specific genes. BRG1 or BAF47 downregulation disrupt both proliferation and differentiation genetic programs expression. Interestingly, BRG1 and BAF47 are part of the SWI/SNF remodeling complex as well as the N-CoR-1 repressor complex in proliferating myoblasts. However, our data show that, upon myogenic differentiation, BAF47 shifts in favor of N-CoR-1 complex. Finally, BRG1 and BAF47 are well-known tumor suppressors but, strikingly, only BAF47 seems essential in the myoblasts irreversible cell cycle exit. Together, our data unravel differential roles for SWI/SNF subunits in muscle differentiation, with BAF47 playing a dual role both in the permanent cell cycle exit and in the regulation of muscle-specific genes.     

TGF-β induces global changes in DNA methylation during the epithelial-to-mesenchymal transition in ovarian cancer cells

A key step in the process of metastasis is the epithelial-to-mesenchymal transition (EMT). We hypothesized that epigenetic mechanisms play a key role in EMT and to test this hypothesis we analyzed global and gene-specific changes in DNA methylation during TGF-β-induced EMT in ovarian cancer cells. Epigenetic profiling using the Infinium HumanMethylation450 BeadChip (HM450) revealed extensive (P < 0.01) methylation changes after TGF-β stimulation (468 and 390 CpG sites altered at 48 and 120 h post cytokine treatment, respectively). The majority of gene-specific TGF-β-induced methylation changes occurred in CpG islands located in or near promoters (193 and 494 genes hypermethylated at 48 and 120 h after TGF-β stimulation, respectively). Furthermore, methylation changes were sustained for the duration of TGF-β treatment and reversible after the cytokine removal. Pathway analysis of the hypermethylated loci identified functional networks strongly associated with EMT and cancer progression, including cellular movement, cell cycle, organ morphology, cellular development, and cell death and survival. Altered methylation and corresponding expression of specific genes during TGF-β-induced EMT included CDH1 (E-cadherin) and COL1A1 (collagen 1A1). Furthermore, TGF-β induced both expression and activity of DNA methyltransferases (DNMT) -1, -3A, and -3B, and treatment with the DNMT inhibitor SGI-110 prevented TGF-β-induced EMT. These results demonstrate that dynamic changes in the DNA methylome are implicated in TGF-β-induced EMT and metastasis. We suggest that targeting DNMTs may inhibit this process by reversing the EMT genes silenced by DNA methylation in cancer.     

Early-Stage Induction of SWI/SNF Mutations during Esophageal Squamous Cell Carcinogenesis

The SWI/SNF chromatin remodeling complex is frequently inactivated by somatic mutations of its various components in various types of cancers, and also by aberrant DNA methylation. However, its somatic mutations and aberrant methylation in esophageal squamous cell carcinomas (ESCCs) have not been fully analyzed. In this study, we aimed to clarify in ESCC, what components of the SWI/SNF complex have somatic mutations and aberrant methylation, and when somatic mutations of the SWI/SNF complex occur. Deep sequencing of components of the SWI/SNF complex using a bench-top next generation sequencer revealed that eight of 92 ESCCs (8.7%) had 11 somatic mutations of 7 genes, ARID1A, ARID2, ATRX, PBRM1, SMARCA4, SMARCAL1, and SMARCC1. The SMARCA4 mutations were located in the Forkhead (85Ser>Leu) and SNF2 family N-terminal (882Glu>Lys) domains. The PBRM1 mutations were located in a bromodomain (80Asn>Ser) and an HMG-box domain (1,377Glu>Lys). For most mutations, their mutant allele frequency was 31–77% (mean 61%) of the fraction of cancer cells in the same samples, indicating that most of the cancer cells in individual ESCC samples had the SWI/SNF mutations on one allele, when present. In addition, a BeadChip array analysis revealed that a component of the SWI/SNF complex, ACTL6B, had aberrant methylation at its promoter CpG island in 18 of 52 ESCCs (34.6%). These results showed that genetic and epigenetic alterations of the SWI/SNF complex are present in ESCCs, and suggested that genetic alterations are induced at an early stage of esophageal squamous cell carcinogenesis.     

SWI/SNF Subunits SMARCA4, SMARCD2 and DPF2 Collaborate in MLL-Rearranged Leukaemia Maintenance

Alterations in chromatin structure caused by deregulated epigenetic mechanisms collaborate with underlying genetic lesions to promote cancer. SMARCA4/BRG1, a core component of the SWI/SNF ATP-dependent chromatin-remodelling complex, has been implicated by its mutational spectrum as exerting a tumour-suppressor function in many solid tumours; recently however, it has been reported to sustain leukaemogenic transformation in MLL-rearranged leukaemia in mice. Here we further explore the role of SMARCA4 and the two SWI/SNF subunits SMARCD2/BAF60B and DPF2/BAF45D in leukaemia. We observed the selective requirement for these proteins for leukaemic cell expansion and self-renewal in-vitro as well as in leukaemia. Gene expression profiling in human cells of each of these three factors suggests that they have overlapping functions in leukaemia. The gene expression changes induced by loss of the three proteins demonstrate that they are required for the expression of haematopoietic stem cell associated genes but in contrast to previous results obtained in mouse cells, the three proteins are not required for the expression of c-MYC regulated genes.     

Differential expression of key subunits of SWI/SNF chromatin remodeling complexes in porcine embryos derived in vitro or in vivo

In vitro embryo production is an established method for both humans and animals, but is fraught with inferior development and health issues in offspring born after in vitro fertilization procedures. Analysis of epigenetic changes caused by exposure to in vitro conditions should shed light on potential sources of these phenotypes. Using immunocytochemistry, we investigated the localization and relative abundance of components associated with the SWI/SNF (Switch/Sucrose non‐fermentable) chromatin‐remodeling complex—including BAF155, BAF170, BAF180, BAF53A, BAF57, BAF60A, BAF45D, ARID1A, ARID1B, ARID2, SNF5, and BRD7—in oocytes and in in vitro‐produced and in vivo‐derived porcine embryos. Differences in the localization of BAF155, BAF170, BAF60A, and ARID1B among these sources indicate that improper timing of chromatin remodeling and cellular differentiation might occur in early preimplantation embryos produced and cultured in vitro.     

Histone deacetylase 2 knockout suppresses immune escape of triple-negative breast cancer cells via downregulating PD-L1 expression

The PD-L1 overexpression is an important event of immune escape and metastasis in triple-negative breast cancer (TNBC), but the molecular mechanism remains to be determined. Interferon gamma (IFNγ) represents a major driving force behind PD-L1 expression in tumor microenvironment, and histone deacetylase 2 (HDAC2) is required for IFN signaling. Here, we investigated the regulation of HDAC2 on the IFNγ-induced PD-L1 expression in TNBC cells. We found the HDAC2 and PD-L1 expression in TNBC was significantly higher than that in non-TNBC, and HDAC2 was positively correlated with PD-L1 expression. HDAC2 promoted PD-L1 induction by upregulating the phosphorylation of JAK1, JAK2, and STAT1, as well as the translocation of STAT1 to the nucleus and the recruitment of STAT1 to the PD-L1 promoter. Meanwhile, HDAC2 was recruited to the PD-L1 promoter by STAT1, and HDAC2 knockout compromised IFNγ-induced upregulation of H3K27, H3K9 acetylation, and the BRD4 recruitment in PD-L1 promoter. In addition, significant inhibition of proliferation, colony formation, migration, and cell cycle of TNBC cells were observed following knockout of HDAC2 in vitro. Furthermore, HDAC2 knockout reduced IFNγ-induced PD-L1 expression, lymphocyte infiltration, and retarded tumor growth and metastasis in the breast cancer mouse models. This study may provide evidence that HDAC2 promotes IFNγ-induced PD-L1 expression, suggesting a way for enhanced antitumor immunity when targeting the HDAC2 in TNBC.Subject terms: Breast cancer, Immune evasion