Dataset Information

TSC1 controls IL-1? expression in macrophages via mTORC1-dependent C/EBP? pathway.

ABSTRACT: The tuberous sclerosis complex 1 (TSC1) is a tumor suppressor that inhibits the mammalian target of rapamycin (mTOR), which serves as a key regulator of inflammatory responses after bacterial stimulation in monocytes, macrophages, and primary dendritic cells. Previous studies have shown that TSC1 knockout (KO) macrophages produced increased inflammatory responses including tumor necrosis factor-? (TNF-?) and IL-12 to pro-inflammatory stimuli, but whether and how TSC1 regulates pro-IL-1? expression remains unclear. Here using a mouse model in which myeloid lineage-specific deletion of TSC1 leads to constitutive mTORC1 activation, we found that TSC1 deficiency resulted in impaired expression of pro-IL-1? in macrophages following lipopolysaccharide stimulation. Such decreased pro-IL-1? expression in TSC1 KO macrophages was rescued by reducing mTORC1 activity with rapamycin or deletion of mTOR. Rictor deficiency has no detectable effect on pro-IL-1? synthesis, suggesting that TSC1 positively controls pro-IL-1? expression through mTORC1 pathway. Moreover, mechanism studies suggest that mTORC1-mediated downregulation of the CCAAT enhancer-binding protein (C/EBP?) critically contributes to the defective pro-IL-1? expression. Overall, these findings highlight a critical role of TSC1 in regulating innate immunity by control of the mTOR1-C/EBP? pathway.

SUBMITTER: Yang T

PROVIDER: S-EPMC5037279 | biostudies-literature | 2016 Sep

REPOSITORIES: biostudies-literature

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TSC1 controls IL-1β expression in macrophages via mTORC1-dependent C/EBPβ pathway.

Yang Tao T Zhu Linnan L Zhai Yanhua Y Zhao Qingjie Q Peng Jianxia J Zhang Hongbing H Yang Zhongzhou Z Zhang Lianfeng L Ding Wenjun W Zhao Yong Y

Cellular & molecular immunology 20150525 5

The tuberous sclerosis complex 1 (TSC1) is a tumor suppressor that inhibits the mammalian target of rapamycin (mTOR), which serves as a key regulator of inflammatory responses after bacterial stimulation in monocytes, macrophages, and primary dendritic cells. Previous studies have shown that TSC1 knockout (KO) macrophages produced increased inflammatory responses including tumor necrosis factor-α (TNF-α) and IL-12 to pro-inflammatory stimuli, but whether and how TSC1 regulates pro-IL-1β expressi ...[more]

PMID: 27593484

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Project description:ObjectiveC/EBPβ, a crucial transcription factor, regulates innate immunity and inflammatory responses. However, the role played by C/EBPβ in alveolar macrophage (AM) inflammatory responses remains unknown. This study aimed to investigate the role and mechanism of C/EBPβ in alveolar macrophages (AMs) from the transcriptional level and to search for natural compounds targeting C/EBPβ.MethodsRat AMs were infected with Lv-sh-C/EBPβ and treated with LPS, and the expression levels of iNOS, TNF-α, IL-6, and IL-1β were measured by RT-qPCR, Western blotting, and ELISA. Mechanistically, transcriptome sequencing (RNA-seq) revealed changes in gene expression patterns in AMs after LPS stimulation and C/EBPβ knockdown. Functional enrichment analyses and rescue experiments identified and validated inflammation-associated cell signaling pathways regulated by C/EBPβ. Furthermore, virtual screening was used to search for natural compounds that inhibit C/EBPβ with the structure of helenalin as a reference.ResultsFollowing stimulation with LPS, AMs exhibited an increased expression of C/EBPβ. C/EBPβ knockdown significantly decreased the expression levels of inflammatory mediators. A total of 374 differentially expressed genes (DEGs) were identified between LPS-stimulated C/EBPβ knockdown and negative control cells. The NOD-like receptor signaling may be a key target for C/EBPβ, according to functional enrichment analyses of the DEGs. Further experiments showed that the muramyl dipeptide (MDP, NOD2 agonist) reversed the downregulation of inflammatory mediators and the NF-κB pathway caused by the C/EBPβ knockdown. The virtual screening revealed that N-caffeoyltryptophan, orilotimod, and petasiphenone have comparable pharmacological properties to helenalin (a known C/EBPβ inhibitor) and demonstrate a great binding capacity to C/EBPβ.ConclusionAblation of C/EBPβ may attenuate LPS-induced inflammatory damage in AMs by inhibiting the NOD2 receptor signaling pathway. Three natural compounds, N-caffeoyltryptophan, orilotimod, and petasiphenone, may be potential C/EBPβ inhibitors.

Project description:BackgroundCCAAT enhancer-binding protein (C/EBP)β regulates gene expression in multiple organ systems and cell types, including astrocytes in the central nervous system (CNS). Inflammatory stimuli, interleukin (IL)-1β, tumor necrosis factor-α, human immunodeficiency virus (HIV)-1 and lipopolysaccharide induce astrocyte C/EBPβ expression. C/EBPβ is detectable in brains of Alzheimer's disease (AD), Parkinson's disease (PD) and HIV-1-associated dementia (HAD) patients, yet little is known about how C/EBPβ contributes to astrocyte gene regulation during neuroinflammation.MethodsThe expression of 92 human inflammation genes was compared between IL-1β-treated primary human astrocytes and astrocytes transfected with C/EBPβ-specific small interfering (si)RNA prior to IL-1β treatment for 12 h. Transcripts altered by>two-fold compared to control were subjected to one-way analysis of variance and Newman-Keuls post-test for multiple comparisons. Expression of two genes, cyclooxygenase-2 (COX-2) and bradykinin receptor B2 (BDKRB2) was further confirmed in additional human astrocyte donors. Astrocytes were treated with mitogen-activated protein kinase-selective inhibitors, then with IL-1β for 12 or 24 h followed by COX-2 and BDKRB2, expression analyses.ResultsIL-1β altered expression of 29 of 92 human inflammation genes by at least two-fold in primary human astrocytes in 12 h. C/EBPβ knockdown affected expression of 17 out of 29 IL-1β-regulated genes by>25%. Two genes relevant to neuroinflammation, COX-2 and BDKRB2, were robustly decreased and increased, respectively, in response to C/EBPβ knockdown, and expression was confirmed in two additional donors. COX-2 and BDKRB2 mRNA remained altered in siRNA-transfected astrocytes at 12, 24 or 72 h. Inhibiting p38 kinase (p38K) activation blocked IL-1β-induced astrocyte COX-2 mRNA and protein expression, but not IL-1β-induced astrocyte BDKRB2 expression. Inhibiting extracellular-regulated kinase (ERK)1/2 activation blocked IL-1β-induced BDKRB2 mRNA expression while increasing COX-2 expression.ConclusionThese data support an essential role for IL-1β in the CNS and identify new C/EBPβ functions in astrocytes. Additionally, this work suggests p38K and ERK1/2 pathways may regulate gene expression in a complementary manner to fine tune the IL-1β-mediated astrocyte inflammatory response. Delineating a role for C/EBPβ and other involved transcription factors in human astrocyte inflammatory response may lead to effective therapies for AD, PD, HAD and other neurological disorders.

Dataset Information

TSC1 controls IL-1? expression in macrophages via mTORC1-dependent C/EBP? pathway.

Publications

TSC1 controls IL-1β expression in macrophages via mTORC1-dependent C/EBPβ pathway.

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