HDAC2 Is Involved in the Regulation of BRN3A in Melanocytes and Melanoma

The neural crest transcription factor BRN3A is essential for the proliferation and survival of melanoma cells. It is frequently expressed in melanoma but not in normal melanocytes or benign nevi. The mechanisms underlying the aberrant expression of BRN3A are unknown. Here, we investigated the epigenetic regulation of BRN3A in melanocytes and melanoma cell lines treated with DNA methyltransferase (DNMT), histone acetyltransferase (HAT), and histone deacetylase (HDAC) inhibitors. DNMT and HAT inhibition did not significantly alter BRN3A expression levels, whereas panHDAC inhibition by trichostatin A led to increased expression. Treatment with the isoform-specific HDAC inhibitor mocetinostat, but not with PCI-34051, also increased BRN3A expression levels, suggesting that class I HDACs HDAC1, HDAC2, and HDAC3, and class IV HDAC11, were involved in the regulation of BRN3A expression. Transient silencing of HDACs 1, 2, 3, and 11 by siRNAs revealed that, specifically, HDAC2 inhibition was able to increase BRN3A expression. ChIP-Seq analysis uncovered that HDAC2 inhibition specifically increased H3K27ac levels at a distal enhancer region of the BRN3A gene. Altogether, our data suggest that HDAC2 is a key epigenetic regulator of BRN3A in melanocytes and melanoma cells. These results highlight the importance of epigenetic mechanisms in regulating melanoma oncogenes.

Beyond HAT Adaptor: TRRAP Liaisons with Sp1-Mediated Transcription

The members of the phosphatidylinositol 3-kinase-related kinase (PIKK) family play vital roles in multiple biological processes, including DNA damage response, metabolism, cell growth, mRNA decay, and transcription. TRRAP, as the only member lacking the enzymatic activity in this family, is an adaptor protein for several histone acetyltransferase (HAT) complexes and a scaffold protein for multiple transcription factors. TRRAP has been demonstrated to regulate various cellular functions in cell cycle progression, cell stemness maintenance and differentiation, as well as neural homeostasis. TRRAP is known to be an important orchestrator of many molecular machineries in gene transcription by modulating the activity of some key transcription factors, including E2F1, c-Myc, p53, and recently, Sp1. This review summarizes the biological and biochemical studies on the action mode of TRRAP together with the transcription factors, focusing on how TRRAP-HAT mediates the transactivation of Sp1-governing biological processes, including neurodegeneration.

PER2 mediates CREB-dependent light induction of the clock gene Per1

Light affects many physiological processes in mammals such as entrainment of the circadian clock, regulation of mood, and relaxation of blood vessels. At the molecular level, a stimulus such as light initiates a cascade of kinases that phosphorylate CREB at various sites, including serine 133 (S133). This modification leads CREB to recruit the co-factor CRCT1 and the histone acetyltransferase CBP to stimulate the transcription of genes containing a CRE element in their promoters, such as Period 1 (Per1). However, the details of this pathway are poorly understood. Here we provide evidence that PER2 acts as a co-factor of CREB to facilitate the formation of a transactivation complex on the CRE element of the Per1 gene regulatory region in response to light or forskolin. Using in vitro and in vivo approaches, we show that PER2 modulates the interaction between CREB and its co-regulator CRTC1 to support complex formation only after a light or forskolin stimulus. Furthermore, the absence of PER2 abolished the interaction between the histone acetyltransferase CBP and CREB. This process was accompanied by a reduction of histone H3 acetylation and decreased recruitment of RNA Pol II to the Per1 gene. Collectively, our data show that PER2 supports the stimulus-dependent induction of the Per1 gene via modulation of the CREB/CRTC1/CBP complex.

RAP1 regulates TIP60 function during fate transition between 2 cell-like and pluripotent states

In mammals, the conserved telomere binding protein RAP1 serves a diverse set of non-telomeric functions including activation of the NF-kB signaling pathway, maintenance of metabolic function in vivo, and transcriptional regulation. Here, we uncover the mechanism by which RAP1 modulates gene expression. Using a separation-of-function allele, we show that RAP1 transcriptional regulation is independent of TRF2-mediated binding to telomeres and does not involve direct binding to genomic loci. Instead, RAP1 interacts with the TIP60/p400 complex and modulates its histone acetyltransferase activity. Notably, we show that deletion of RAP1 in mouse embryonic stem cells increases the fraction of 2-cell-like cells. Specifically, RAP1 enhances the repressive activity of Tip60/p400 across a subset of 2-cell-stage genes, including Zscan4 and the endogenous retrovirus MERVL. Preferential upregulation of genes proximal to MERVL elements in Rap1 deficient settings implicate these endogenous retroviral elements in the de-repression of proximal genes. Altogether, our study reveals an unprecedented link between RAP1 and TIP60/p400 complex in the regulation of totipotency.

Inhibition of histone acetyltransferase function radiosensitizes CREBBP/EP300 mutants via repression of homologous recombination, potentially targeting a gain of function

Despite radiation forming the curative backbone of over 50% of malignancies, there are no genomically-driven radiosensitizers for clinical use. Herein we perform in vivo shRNA screening to identify targets generally associated with radiation response as well as those exhibiting a genomic dependency. This identifies the histone acetyltransferases CREBBP/EP300 as a target for radiosensitization in combination with radiation in cognate mutant tumors. Further in vitro and in vivo studies confirm this phenomenon to be due to repression of homologous recombination following DNA damage and reproducible using chemical inhibition of histone acetyltransferase (HAT), but not bromodomain function. Selected mutations in CREBBP lead to a hyperacetylated state that increases CBP and BRCA1 acetylation, representing a gain of function targeted by HAT inhibition. Additionally, mutations in CREBBP/EP300 are associated with recurrence following radiation in squamous cell carcinoma cohorts. These findings provide both a mechanism of resistance and the potential for genomically-driven treatment.

Double peroxidase and histone acetyltransferase AgTip60 maintain innate immune memory in primed mosquitoes

Immune priming in Anopheles gambiae is mediated by the systemic release of a hemocyte differentiation factor (HDF), a complex of lipoxin A4 bound to Evokin, a lipid carrier. HDF increases the proportion of circulating granulocytes and enhances mosquito cellular immunity. Here, we show that Evokin is present in hemocytes and fat-body cells, and messenger RNA (mRNA) expression increases significantly after immune priming. The double peroxidase (DBLOX) enzyme, present in insects but not in vertebrates, is essential for HDF synthesis. DBLOX is highly expressed in oenocytes in the fat-body tissue, and these cells increase in number in primed mosquitoes. We provide direct evidence that the histone acetyltransferase AgTip60 (AGAP001539) is also essential for a sustained increase in oenocyte numbers, HDF synthesis, and immune priming. We propose that oenocytes may function as a population of cells that are reprogrammed, and orchestrate and maintain a broad, systemic, and long-lasting state of enhanced immune surveillance in primed mosquitoes.

Zika virus infection drives epigenetic modulation of immunity by the histone acetyltransferase CBP of Aedes aegypti

Epigenetic mechanisms are responsible for a wide range of biological phenomena in insects, controlling embryonic development, growth, aging and nutrition. Despite this, the role of epigenetics in shaping insect-pathogen interactions has received little attention. Gene expression in eukaryotes is regulated by histone acetylation/deacetylation, an epigenetic process mediated by histone acetyltransferases (HATs) and histone deacetylases (HDACs). In this study, we explored the role of the Aedes aegypti histone acetyltransferase CBP (AaCBP) after infection with Zika virus, focusing on the two main immune tissues, the midgut and fat body. We showed that the expression and activity of AaCBP could be positively modulated by blood meal and Zika infection. Nevertheless, Zika-infected mosquitoes that were silenced for AaCBP revealed a significant reduction in the acetylation of H3K27 (CBP target-marker), followed by downmodulation of the expression of immune genes, higher titers of Zika virus and lower survival rates. Importantly, in Zika-infected mosquitoes that were treated with sodium butyrate, a histone deacetylase inhibitor, their capacity to fight virus infection was rescued. Our data point to a direct correlation among histone hyperacetylation by AaCBP, upregulation of antimicrobial peptide genes and increased survival of Zika-infected-A. aegypti.