The IRG1-itaconate axis protects from cholesterol-induced inflammation and atherosclerosis.

Atherosclerosis is fueled by a failure to resolve lipid-driven inflammation within the vasculature that drives plaque formation. Therapeutic approaches to reverse atherosclerotic inflammation are needed to address the rising global burden of cardiovascular disease (CVD). Recently, metabolites have gained attention for their immunomodulatory properties, including itaconate, which is generated from the tricarboxylic acid-intermediate cis-aconitate by the enzyme Immune Responsive Gene 1 (IRG1/ACOD1). Here, we tested the therapeutic potential of the IRG1-itaconate axis for human atherosclerosis. Using single-cell RNA sequencing (scRNA-seq), we found that IRG1 is up-regulated in human coronary atherosclerotic lesions compared to patient-matched healthy vasculature, and in mouse models of atherosclerosis, where it is primarily expressed by plaque monocytes, macrophages, and neutrophils. Global or hematopoietic Irg1-deficiency in mice increases atherosclerosis burden, plaque macrophage and lipid content, and expression of the proatherosclerotic cytokine interleukin (IL)-1ß. Mechanistically, absence of Irg1 increased macrophage lipid accumulation, and accelerated inflammation via increased neutrophil extracellular trap (NET) formation and NET-priming of the NLRP3-inflammasome in macrophages, resulting in increased IL-1ß release. Conversely, supplementation of the Irg1-itaconate axis using 4-octyl itaconate (4-OI) beneficially remodeled advanced plaques and reduced lesional IL-1ß levels in mice. To investigate the effects of 4-OI in humans, we leveraged an ex vivo systems-immunology approach for CVD drug discovery. Using CyTOF and scRNA-seq of peripheral blood mononuclear cells treated with plasma from CVD patients, we showed that 4-OI attenuates proinflammatory phospho-signaling and mediates anti-inflammatory rewiring of macrophage populations. Our data highlight the relevance of pursuing IRG1-itaconate axis supplementation as a therapeutic approach for atherosclerosis in humans.

An epigenetic switch controls an alternative NR2F2 isoform that unleashes a metastatic program in melanoma.

Metastatic melanoma develops once transformed melanocytic cells begin to de-differentiate into migratory and invasive melanoma cells with neural crest cell (NCC)-like and epithelial-to-mesenchymal transition (EMT)-like features. However, it is still unclear how transformed melanocytes assume a metastatic melanoma cell state. Here, we define DNA methylation changes that accompany metastatic progression in melanoma patients and discover Nuclear Receptor Subfamily 2 Group F, Member 2 - isoform 2 (NR2F2-Iso2) as an epigenetically regulated metastasis driver. NR2F2-Iso2 is transcribed from an alternative transcriptional start site (TSS) and it is truncated at the N-terminal end which encodes the NR2F2 DNA-binding domain. We find that NR2F2-Iso2 expression is turned off by DNA methylation when NCCs differentiate into melanocytes. Conversely, this process is reversed during metastatic melanoma progression, when NR2F2-Iso2 becomes increasingly hypomethylated and re-expressed. Our functional and molecular studies suggest that NR2F2-Iso2 drives metastatic melanoma progression by modulating the activity of full-length NR2F2 (Isoform 1) over EMT- and NCC-associated target genes. Our findings indicate that DNA methylation changes play a crucial role during metastatic melanoma progression, and their control of NR2F2 activity allows transformed melanocytes to acquire NCC-like and EMT-like features. This epigenetically regulated transcriptional plasticity facilitates cell state transitions and metastatic spread.

Melanoma-Secreted Amyloid Beta Suppresses Neuroinflammation and Promotes Brain Metastasis.

Brain metastasis is a significant cause of morbidity and mortality in multiple cancer types and represents an unmet clinical need. The mechanisms that mediate metastatic cancer growth in the brain parenchyma are largely unknown. Melanoma, which has the highest rate of brain metastasis among common cancer types, is an ideal model to study how cancer cells adapt to the brain parenchyma. Our unbiased proteomics analysis of melanoma short-term cultures revealed that proteins implicated in neurodegenerative pathologies are differentially expressed in melanoma cells explanted from brain metastases compared with those derived from extracranial metastases. We showed that melanoma cells require amyloid beta (Aß) for growth and survival in the brain parenchyma. Melanoma-secreted Aß activates surrounding astrocytes to a prometastatic, anti-inflammatory phenotype and prevents phagocytosis of melanoma by microglia. Finally, we demonstrate that pharmacologic inhibition of Aß decreases brain metastatic burden. SIGNIFICANCE: Our results reveal a novel mechanistic connection between brain metastasis and Alzheimer's disease, two previously unrelated pathologies; establish Aß as a promising therapeutic target for brain metastasis; and demonstrate suppression of neuroinflammation as a critical feature of metastatic adaptation to the brain parenchyma. This article is highlighted in the In This Issue feature, p. 1171.

The histone demethylase PHF8 regulates TGFß signaling and promotes melanoma metastasis.

The contribution of epigenetic dysregulation to metastasis remains understudied. Through a meta-analysis of gene expression datasets followed by a mini-screen, we identified Plant Homeodomain Finger protein 8 (PHF8), a histone demethylase of the Jumonji C protein family, as a previously unidentified prometastatic gene in melanoma. Loss- and gain-of-function approaches demonstrate that PHF8 promotes cell invasion without affecting proliferation in vitro and increases dissemination but not subcutaneous tumor growth in vivo, thus supporting its specific contribution to the acquisition of metastatic potential. PHF8 requires its histone demethylase activity to enhance melanoma cell invasion. Transcriptomic and epigenomic analyses revealed that PHF8 orchestrates a molecular program that directly controls the TGFß signaling pathway and, as a consequence, melanoma invasion and metastasis. Our findings bring a mechanistic understanding of epigenetic regulation of metastatic fitness in cancer, which may pave the way for improved therapeutic interventions.

Network models of primary melanoma microenvironments identify key melanoma regulators underlying prognosis.

Melanoma is the most lethal skin malignancy, driven by genetic and epigenetic alterations in the complex tumour microenvironment. While large-scale molecular profiling of melanoma has identified molecular signatures associated with melanoma progression, comprehensive systems-level modeling remains elusive. This study builds up predictive gene network models of molecular alterations in primary melanoma by integrating large-scale bulk-based multi-omic and single-cell transcriptomic data. Incorporating clinical, epigenetic, and proteomic data into these networks reveals key subnetworks, cell types, and regulators underlying melanoma progression. Tumors with high immune infiltrates are found to be associated with good prognosis, presumably due to induced CD8+ T-cell cytotoxicity, via MYO1F-mediated M1-polarization of macrophages. Seventeen key drivers of the gene subnetworks associated with poor prognosis, including the transcription factor ZNF180, are tested for their pro-tumorigenic effects in vitro. The anti-tumor effect of silencing ZNF180 is further validated using in vivo xenografts. Experimentally validated targets of ZNF180 are enriched in the ZNF180 centered network and the known pathways such as melanoma cell maintenance and immune cell infiltration. The transcriptional networks and their critical regulators provide insights into the molecular mechanisms of melanomagenesis and pave the way for developing therapeutic strategies for melanoma.

Diminished Fear Extinction in Adolescents Is Associated With an Altered Somatostatin Interneuron-Mediated Inhibition in the Infralimbic Cortex.

BACKGROUND: Rodents and humans show an attenuation of fear extinction during adolescence, which coincides with the onset of several psychiatric disorders. Although the ethological relevance and the underlying mechanism are largely unknown, the suppression of fear extinction during adolescence is associated with a diminished plasticity in the glutamatergic neurons of the infralimbic medial prefrontal cortex, a brain region critical for fear extinction. Given the putative effect of synaptic inhibition on glutamatergic neuron activity, we studied whether gamma-aminobutyric acidergic neurons in the infralimbic medial prefrontal cortex are involved in the suppression of fear extinction during adolescence. METHODS: We assessed membrane and synaptic properties in parvalbumin-positive interneurons (PVINs) and somatostatin-positive interneurons (SSTINs) in male preadolescent, adolescent, and adult mice. The effect of fear conditioning and extinction on PVIN-pyramidal neuron and SSTIN-pyramidal neuron synapses in male preadolescent, adolescent, and adult mice was evaluated using an optogenetic approach. RESULTS: The development of the membrane excitability of PVINs is delayed and reaches maturity only by adulthood, while the SSTIN membrane properties are developed early and remain stable during development from preadolescence to adulthood. Although the synaptic inhibition mediated by PVINs undergoes a protracted development, it does not exhibit a fear behavior-specific plasticity. However, the synaptic inhibition mediated by SSTINs undergoes an adolescence-specific enhancement, and this increased inhibition is suppressed by fear learning but is not restored by extinction training. This altered plasticity during adolescence overlapped with a reduction in calcium-permeable glutamate receptors in SSTINs. CONCLUSIONS: The adolescence-specific plasticity in the SSTINs might play a role in fear extinction suppression during adolescence in mice.

Opposing effects of an atypical glycinergic and substance P transmission on interpeduncular nucleus plasticity.

The medial habenula-interpeduncular nucleus (MHb-IPN) pathway has recently been implicated in the suppression of fear memory. A notable feature of this pathway is the corelease of neurotransmitters and neuropeptides from MHb neurons. Our studies in mice reveal that an activation of substance P-positive dorsomedial habenula (dMHb) neurons results in simultaneous release of glutamate and glycine in the lateral interpeduncular nucleus (LIPN). This glycine receptor activity inhibits an activity-dependent long-lasting potentiation of glutamatergic synapses in LIPN neurons, while substance P enhances this plasticity. An endocannabinoid CB1 receptor-mediated suppression of GABAB receptor activity allows substance P to induce a long-lasting increase in glutamate release in LIPN neurons. Consistent with the substance P-dependent synaptic potentiation in the LIPN, the NK1R in the IPN is involved in fear extinction but not fear conditioning. Thus, our study describes a novel plasticity mechanism in the LIPN and a region-specific role of substance P in fear extinction.