RESUMO
Neuronal activity causes the rapid expression of immediate early genes that are crucial for experience-driven changes to synapses, learning, and memory. Here, using both molecular and genome-wide next-generation sequencing methods, we report that neuronal activity stimulation triggers the formation of DNA double strand breaks (DSBs) in the promoters of a subset of early-response genes, including Fos, Npas4, and Egr1. Generation of targeted DNA DSBs within Fos and Npas4 promoters is sufficient to induce their expression even in the absence of an external stimulus. Activity-dependent DSB formation is likely mediated by the type II topoisomerase, Topoisomerase IIß (Topo IIß), and knockdown of Topo IIß attenuates both DSB formation and early-response gene expression following neuronal stimulation. Our results suggest that DSB formation is a physiological event that rapidly resolves topological constraints to early-response gene expression in neurons.
Assuntos
Quebras de DNA de Cadeia Dupla , Neurônios/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fator de Ligação a CCCTC , DNA Topoisomerases Tipo II/análise , DNA Topoisomerases Tipo II/metabolismo , Proteínas de Ligação a DNA/análise , Proteínas de Ligação a DNA/metabolismo , Proteína 1 de Resposta de Crescimento Precoce/genética , Etoposídeo/farmacologia , Regulação da Expressão Gênica , Genes fos , Estudo de Associação Genômica Ampla , Camundongos , Proteínas Repressoras/metabolismo , Transcriptoma/efeitos dos fármacosRESUMO
Regulatory B (Breg) cells have been shown to play a critical role in immune homeostasis and in autoimmunity models. We have recently demonstrated that combined anti-T cell immunoglobulin domain and mucin domain-1 and anti-CD45RB antibody treatment results in tolerance to full MHC-mismatched islet allografts in mice by generating Breg cells that are necessary for tolerance. Breg cells are antigen-specific and are capable of transferring tolerance to untreated, transplanted animals. Here, we demonstrate that adoptively transferred Breg cells require the presence of regulatory T (Treg) cells to establish tolerance, and that adoptive transfer of Breg cells increases the number of Treg cells. Interaction with Breg cells in vivo induces significantly more Foxp3 expression in CD4(+) CD25(-) T cells than with naive B cells. We also show that Breg cells express the TGF-ß associated latency-associated peptide and that Breg-cell mediated graft prolongation post-adoptive transfer is abrogated by neutralization of TGF-ß activity. Breg cells, like Treg cells, demonstrate preferential expression of both C-C chemokine receptor 6 and CXCR3. Collectively, these findings suggest that in this model of antibody-induced transplantation tolerance, Breg cells promote graft survival by promoting Treg-cell development, possibly via TGF-ß production.