ABSTRACT
Gasdermin B (GSDMB) belongs to a large family of pore-forming cytolysins that execute inflammatory cell death programs. While genetic studies have linked GSDMB polymorphisms to human disease, its function in the immunological response to pathogens remains poorly understood. Here, we report a dynamic host-pathogen conflict between GSDMB and the IpaH7.8 effector protein secreted by enteroinvasive Shigella flexneri. We show that IpaH7.8 ubiquitinates and targets GSDMB for 26S proteasome destruction. This virulence strategy protects Shigella from the bacteriocidic activity of natural killer cells by suppressing granzyme-A-mediated activation of GSDMB. In contrast to the canonical function of most gasdermin family members, GSDMB does not inhibit Shigella by lysing host cells. Rather, it exhibits direct microbiocidal activity through recognition of phospholipids found on Gram-negative bacterial membranes. These findings place GSDMB as a central executioner of intracellular bacterial killing and reveal a mechanism employed by pathogens to counteract this host defense system.
Subject(s)
Biomarkers, Tumor/metabolism , Host-Pathogen Interactions , Killer Cells, Natural/immunology , Neoplasm Proteins/metabolism , Pore Forming Cytotoxic Proteins/metabolism , Shigella flexneri/physiology , Ubiquitination , Animals , Bacterial Proteins/metabolism , Cardiolipins/metabolism , Cell Line , Cell Membrane/metabolism , Female , Granzymes/metabolism , Humans , Lipid A/metabolism , Mice, Inbred C57BL , Mice, Transgenic , Microbial Viability , Proteasome Endopeptidase Complex/metabolism , Protein Binding , Proteolysis , Substrate SpecificityABSTRACT
MicroRNA cluster mirn23a has previously been shown to promote myeloid development at the expense of lymphoid development in overexpression and knockout mouse models. This polarization is observed early in hematopoietic development, with an increase in common lymphoid progenitors (CLPs) and a decrease in all myeloid progenitor subsets in adult bone marrow. The pool size of multipotential progenitors (MPPs) is unchanged; however, in this report we observe by flow cytometry that polarized subsets of MPPs are changed in the absence of mirn23a. Additionally, in vitro culture of MPPs and sorted MPP transplants showed that these cells have decreased myeloid and increased lymphoid potential in vitro and in vivo. We investigated the mechanism by which mirn23a regulates hematopoietic differentiation and observed that mirn23a promotes myeloid development of hematopoietic progenitors through regulation of hematopoietic transcription factors and signaling pathways. Early transcription factors that direct the commitment of MPPs to CLPs (Ikzf1, Runx1, Satb1, Bach1 and Bach2) are increased in the absence of mirn23a miRNAs as well as factors that commit the CLP to the B cell lineage (FoxO1, Ebf1, and Pax5). Mirn23a appears to buffer transcription factor levels so that they do not stochastically reach a threshold level to direct differentiation. Intriguingly, mirn23a also inversely regulates the PI3 kinase (PI3K)/Akt and BMP/Smad signaling pathways. Pharmacological inhibitor studies, coupled with dominant active/dominant negative biochemical experiments, show that both signaling pathways are critical to mirn23a's regulation of hematopoietic differentiation. Lastly, consistent with mirn23a being a physiological inhibitor of B cell development, we observed that the essential B cell transcription factor EBF1 represses expression of mirn23a. In summary, our data demonstrates that mirn23a regulates a complex array of transcription and signaling pathways to modulate adult hematopoiesis.
Subject(s)
Hematopoiesis/genetics , MicroRNAs/genetics , Animals , Apoptosis , Apoptosis Regulatory Proteins , B-Lymphocytes/metabolism , Carrier Proteins/genetics , Carrier Proteins/metabolism , Cell Cycle Proteins/genetics , Cell Cycle Proteins/metabolism , Cell Differentiation , Cell Line , Cell Proliferation , Down-Regulation , Forkhead Box Protein O1/genetics , Forkhead Box Protein O1/metabolism , Guanylate Kinases/genetics , Guanylate Kinases/metabolism , Membrane Proteins/genetics , Membrane Proteins/metabolism , Mice , Mice, Inbred C57BL , MicroRNAs/metabolism , Mitochondrial Proteins/genetics , Mitochondrial Proteins/metabolism , NIH 3T3 Cells , Phosphatidylinositol 3-Kinases/genetics , Phosphatidylinositol 3-Kinases/metabolism , Promoter Regions, Genetic , Signal Transduction , Trans-Activators/genetics , Trans-Activators/metabolism , Up-RegulationABSTRACT
Research on engagement within community-based psychiatric services in the UK has mainly focussed on factors related to those 'at risk' of non-attendance or non-compliance, with the tacit assumption that those in regular attendance are largely content and hence not a priority. The present study systematically explored the experiences and views of 25 people with severe and enduring mental illness who had regularly attended out-patient settings for more than 5 years. Regular attendance at consultations was not synonymous with satisfaction-in fact it masked varying levels of unmet needs and 'de-humanisation'. In order to establish and maintain non-coercive community services that prioritise 'recovery' above illness and 'risk' containment, it is essential that the experiences of people in established and apparently 'less troublesome' therapeutic relationships are also taken into account and integrated into policy and practice.
