The early macrophage response to pathogens requires dynamic regulation of the nuclear paraspeckle

ABSTRACT To ensure a robust immune response to pathogens without risking immunopathology, the kinetics and amplitude of inflammatory gene expression in macrophages needs to be exquisitely well-controlled. There is a growing appreciation for stress-responsive membraneless organelles (MLOs) regulating various steps of eukaryotic gene expression in response to extrinsic cues. Here, we implicate the nuclear paraspeckle, a highly ordered biomolecular condensate that nucleates on the Neat1 lncRNA, in tuning innate immune gene expression in murine macrophages. In response to a variety of innate agonists, macrophage paraspeckles rapidly aggregate (0.5 h post-stimulation) and disaggregate (2h post-stimulation). Paraspeckle maintenance and aggregation require active transcription and MAPK signaling whereas paraspeckle disaggregation requires degradation of Neat1 via the nuclear RNA exosome. Expression of a large cohort of cytokines, chemokines, and antimicrobial mediators is compromised in lipopolysaccharide-treated macrophages lacking Neat1 , resulting in a failure to express a cohort of pro-inflammatory cytokines, chemokines, and antimicrobial mediators. Consequently, Neat1 KO macrophages cannot control replication of Salmonella enterica serovar Typhimurium or vesicular stomatitis virus. These findings highlight a prominent role for MLOs in orchestrating the macrophage response to pathogens and support a model whereby dynamic assembly and disassembly of paraspeckles reprograms the nuclear RNA binding protein landscape to enable inflammatory gene expression following innate stimuli. SIGNIFICANCE STATEMENT To mount appropriate immune responses and fight infection, macrophages need to sense and respond to pathogen-associated signals with incredible precision. Membraneless organelles (MLOs) are complexes of RNAs and proteins that change in size, shape, and abundance in response to extracellular signals. We hypothesized that an MLO called the nuclear paraspeckle helps macrophages initiate and calibrate innate immune gene expression during infection. We found that paraspeckles rapidly aggregate and then dissolve in macrophages following pathogen sensing. Macrophages lacking paraspeckles cannot properly induce inflammatory genes, resulting in a failure to control replication of intracellular bacterial and viral pathogens. These data suggest that altered paraspeckle dynamics may dysregulate inflammatory gene expression in a variety of human diseases.