RESUMO
Transcriptional programming of the innate immune response is pivotal for host protection. However, the transcriptional mechanisms that link pathogen sensing with innate activation remain poorly understood. During HIV-1 infection, human dendritic cells (DCs) can detect the virus through an innate sensing pathway, leading to antiviral interferon and DC maturation. Here, we develop an iterative experimental and computational approach to map the HIV-1 innate response circuitry in monocyte-derived DCs (MDDCs). By integrating genome-wide chromatin accessibility with expression kinetics, we infer a gene regulatory network that links 542 transcription factors with 21,862 target genes. We observe that an interferon response is required, yet insufficient, to drive MDDC maturation and identify PRDM1 and RARA as essential regulators of the interferon response and MDDC maturation, respectively. Our work provides a resource for interrogation of regulators of HIV replication and innate immunity, highlighting complexity and cooperativity in the regulatory circuit controlling the response to infection.
Assuntos
Células Dendríticas/metabolismo , Redes Reguladoras de Genes , HIV-1/imunologia , Imunidade Inata/genética , Monócitos/metabolismo , Diferenciação Celular , Cromatina/metabolismo , Células Dendríticas/virologia , Feminino , Regulação da Expressão Gênica , Células HEK293 , Infecções por HIV/imunologia , Infecções por HIV/virologia , Humanos , Interferon Tipo I/metabolismo , Masculino , Monócitos/virologia , Regiões Promotoras Genéticas/genética , Receptor alfa de Ácido Retinoico/metabolismo , Fatores de Transcrição/metabolismo , Transcriptoma/genéticaRESUMO
Myeloid dendritic cells (DCs) have the innate capacity to sense pathogens and orchestrate immune responses. However, DCs do not mount efficient immune responses to HIV-1, primarily due to restriction of virus reverse transcription, which prevents accumulation of viral cDNA and limits its detection through the cGAS-STING pathway. By allowing reverse transcription to proceed, we find that DCs detect HIV-1 in distinct phases, before and after virus integration. Blocking integration suppresses, but does not abolish, activation of the transcription factor IRF3, downstream interferon (IFN) responses, and DC maturation. Consistent with two stages of detection, HIV-1 "primes" chromatin accessibility of innate immune genes before and after integration. Once primed, robust IFN responses can be unmasked by agonists of the innate adaptor protein, MyD88, through a process that requires cGAS, STING, IRF3, and nuclear factor κB. Thus, HIV-1 replication increases material available for sensing, and discrete inflammatory inputs tune cGAS signaling to drive DC maturation.
Assuntos
Cromatina/metabolismo , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , HIV-1/imunologia , Interações Hospedeiro-Patógeno/imunologia , Interferons/metabolismo , Linhagem Celular , Feminino , Células HEK293 , Infecções por HIV/metabolismo , HIV-1/patogenicidade , Humanos , Imunidade Inata/genética , Imunidade Inata/imunologia , Fator Regulador 3 de Interferon/metabolismo , Interferon Tipo I/metabolismo , Masculino , Proteínas de Membrana/metabolismo , Fator 88 de Diferenciação Mieloide/metabolismo , NF-kappa B/metabolismo , Nucleotidiltransferases/metabolismo , Transcrição Reversa , Transdução de Sinais , Células THP-1 , Integração Viral , Replicação ViralRESUMO
Cyclospora cayetanensis, a protozoan of emerging concern, causes self-limiting gastroenteritis in immune-competent hosts. It has been established that sequence variability exists in the first internal transcribed spacer region (ITS-1) of the ribosomal DNA operon from collections of oocysts obtained from individual or pooled fecal samples. To determine if single oocysts also exhibited ITS-1 sequence variability, DNA was extracted from individually flow-cytometry-counted oocysts. We determined that ITS-1 sequence variability exists at an individual-genome level for C. cayetanensis and approached or exceeded the variability exhibited among oocyst collections. ITS-1 variability, at the genome level, reduces this region's utility for inferring relationships between strains.
Assuntos
Cyclospora/genética , Ciclosporíase/parasitologia , DNA Espaçador Ribossômico/química , Gastroenterite/parasitologia , Sequência Conservada , Cyclospora/classificação , DNA de Protozoário/química , Fezes/parasitologia , Citometria de Fluxo , Humanos , Oocistos , Filogenia , Reação em Cadeia da Polimerase , Alinhamento de SequênciaRESUMO
Cyclospora cayetanensis is the causative agent of cyclosporiasis, an emerging infectious disease. We present a new method for the purification of C. cayetanensis oocysts from feces using a modified detachment solution and Renocal-sucrose gradient sedimentation. This method yields oocysts free from adherent fecal debris and amenable to processing using flow cytometry.
