RESUMO
During development, inflammation or tissue injury, macrophages may successively engulf and process multiple apoptotic corpses via efferocytosis to achieve tissue homeostasis1. How macrophages may rapidly adapt their transcription to achieve continuous corpse uptake is incompletely understood. Transcriptional pause/release is an evolutionarily conserved mechanism, in which RNA polymerase (Pol) II initiates transcription for 20-60 nucleotides, is paused for minutes to hours and is then released to make full-length mRNA2. Here we show that macrophages, within minutes of corpse encounter, use transcriptional pause/release to unleash a rapid transcriptional response. For human and mouse macrophages, the Pol II pause/release was required for continuous efferocytosis in vitro and in vivo. Interestingly, blocking Pol II pause/release did not impede Fc receptor-mediated phagocytosis, yeast uptake or bacterial phagocytosis. Integration of data from three genomic approaches-precision nuclear run-on sequencing, RNA sequencing, and assay for transposase-accessible chromatin using sequencing (ATAC-seq)-on efferocytic macrophages at different time points revealed that Pol II pause/release controls expression of select transcription factors and downstream target genes. Mechanistic studies on transcription factor EGR3, prominently regulated by pause/release, uncovered EGR3-related reprogramming of other macrophage genes involved in cytoskeleton and corpse processing. Using lysosomal probes and a new genetic fluorescent reporter, we identify a role for pause/release in phagosome acidification during efferocytosis. Furthermore, microglia from egr3-deficient zebrafish embryos displayed reduced phagocytosis of apoptotic neurons and fewer maturing phagosomes, supporting defective corpse processing. Collectively, these data indicate that macrophages use Pol II pause/release as a mechanism to rapidly alter their transcriptional programs for efficient processing of the ingested apoptotic corpses and for successive efferocytosis.
Assuntos
Eferocitose , Macrófagos , RNA Polimerase II , Elongação da Transcrição Genética , Animais , Humanos , Masculino , Camundongos , Apoptose , Citoesqueleto/metabolismo , Proteína 3 de Resposta de Crescimento Precoce/deficiência , Proteína 3 de Resposta de Crescimento Precoce/genética , Eferocitose/genética , Concentração de Íons de Hidrogênio , Macrófagos/imunologia , Macrófagos/metabolismo , Neurônios/metabolismo , Fagossomos/metabolismo , RNA Polimerase II/metabolismo , Fatores de Transcrição/genética , Peixe-Zebra/embriologia , Peixe-Zebra/genética , Fatores de TempoRESUMO
Identification of pro-metastatic genomic alterations is urgently needed to help understand and prevent the fatal course of prostate cancer. Here, we found that the transcription factor EGR3, located at chromosome 8p21.3, is a critical metastasis suppressor. Aberrant deletion of EGR3 was found in up to 59.76% (deep deletions, 16.87%; shallow deletions, 42.89%) of prostate cancer patients. In informatics analysis, EGR3 loss was associated with prostate cancer progression and low survival rates. EGR3 expression inversely correlated with the expressions of epithelial-to-mesenchymal transition (EMT) and metastasis-related gene sets in prostate cancer tissues. In prostate cancer cells, EGR3 blocked the EMT process and suppressed cell migration and invasion. In a mouse model for cancer metastasis, EGR3 overexpression significantly suppressed bone metastases of PC3 and 22Rv1 prostate cancer cells. Mechanistically, EGR3 transcriptionally activated ZFP36, GADD45B, and SOCS3 genes by directly binding to their promoter regions. The EMT-inhibitory and tumor-suppressive roles of the EGR3 downstream genes were identified through in vitro and in silico analyses. Together, our results showed that EGR3 may be a biomarker to predict clinical outcomes and that it plays an important role in the metastatic progression of prostate cancer.
Assuntos
Proteína 3 de Resposta de Crescimento Precoce/deficiência , Predisposição Genética para Doença , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Animais , Linhagem Celular Tumoral , Movimento Celular , Variações do Número de Cópias de DNA , Modelos Animais de Doenças , Progressão da Doença , Transição Epitelial-Mesenquimal/genética , Seguimentos , Deleção de Genes , Regulação Neoplásica da Expressão Gênica , Estudos de Associação Genética , Humanos , Masculino , Camundongos , Metástase Neoplásica , Prognóstico , Modelos de Riscos Proporcionais , Neoplasias da Próstata/mortalidade , Neoplasias da Próstata/patologia , Fatores de Risco , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
TCR-induced NF-AT activation leads to the up-regulation of multiple genes involved in T cell anergy. Since NF-AT is also involved in T cell activation, we have endeavored to dissect TCR-induced activating and inhibitory genetic programs. This approach revealed roles for the early growth response (Egr) family of transcription factors and the Egr coactivator/corepressor NGFI-A-binding protein (NAB)2 in regulating T cell function. TCR-induced Egr-1 and NAB2 enhance T cell function, while Egr-2 and Egr-3 inhibit T cell function. In this report, we demonstrate that Egr-2 and Egr-3 are induced by NF-AT in the absence of AP-1, while Egr-1 and NAB2 both require AP-1-mediated transcription. Our data suggest that Egr-3 is upstream of Egr-2, and that mechanistically Egr-2 and Egr-3 suppress Egr-1 and NAB2 expression. Functionally, T cells from Egr-2 and Egr-3 null mice are hyperresponsive while T cells from Egr-3 transgenic, overexpressing mice are hyporesponsive. Furthermore, an in vivo model of autoimmune pneumonitis reveals that T cells from Egr-3 null mice hasten death while Egr-3-overexpressing T cells cause less disease. Overall, our data suggest that just as the Egr/NAB network of genes control cell fate in other systems, TCR-induced Egr-1, 2, 3 and NAB2 control the fate of antigen recognition in T cells.