Single-Cell Imaging of Caspase-1 Dynamics Reveals an All-or-None Inflammasome Signaling Response

Liu, Ting; Yamaguchi, Yoshifumi; Shirasaki, Yoshitaka; Shikada, Koichi; Yamagishi, Mai; Hoshino, Katsuaki; Kaisho, Tsuneyasu; Takemoto, Kiwamu; Suzuki, Toshihiko; Kuranaga, Erina; Ohara, Osamu; Miura, Masayuki

doi:10.1016/j.celrep.2014.07.012

Cited by 133 publications

(107 citation statements)

References 38 publications

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“…2C). Recent single-cell analysis of caspase-1 dynamics revealed that dead macrophages are the main source of IL-1b secretion within cell populations (23). In contrast to MDM, U937 cells have a prolonged increase in IL-1 gene transcription in response to LPS with a gradual accumulation of cytosolic IL-1 gene products, including IL-1b (24), which explains the high IL-1b secretion despite reduced cell death.…”

Section: Resultsmentioning

confidence: 99%

Cutting Edge: Inflammasome Activation in Primary Human Macrophages Is Dependent on Flagellin

Kortmann

Brubaker

Monack

2015

The Journal of Immunology

146

120

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Murine NLR family, apoptosis inhibitory protein (Naip)1, Naip2, and Naip5/6 are host sensors that detect the cytosolic presence of needle and rod proteins from bacterial type III secretion systems and flagellin, respectively. Previous studies using human-derived macrophage-like cell lines indicate that human macrophages sense the cytosolic needle protein, but not bacterial flagellin. In this study, we show that primary human macrophages readily sense cytosolic flagellin. Infection of primary human macrophages with Salmonella elicits robust cell death and IL-1β secretion that is dependent on flagellin. We show that flagellin detection requires a full-length isoform of human Naip. This full-length Naip isoform is robustly expressed in primary macrophages from healthy human donors, but it is drastically reduced in monocytic tumor cells, THP-1, and U937, rendering them insensitive to cytosolic flagellin. However, ectopic expression of full-length Naip rescues the ability of U937 cells to sense flagellin. In conclusion, human Naip functions to activate the inflammasome in response to flagellin, similar to murine Naip5/6.

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Section: Resultsmentioning

confidence: 99%

Cutting Edge: Inflammasome Activation in Primary Human Macrophages Is Dependent on Flagellin

Kortmann

Brubaker

Monack

2015

The Journal of Immunology

146

120

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“…Although some authors detected IL-1␤ release independent from pyroptosis (31) or pyroptosis without IL-1␤ release (32), others saw pyroptosis as the result of variable inflammasome activation (20,33). A third hypothesis claims an all-or-nothingmechanism with two states: no inflammation or pro-inflammatory cell death/pyroptosis (34). Last, a dual function of caspase-1 was suggested, controlling cell death and autoprocessing with pro-IL-1␤ cleavage through two different pathways (35), and ASC speck formation was found to be essential for release of mature IL-1␤ but not for induction of pyroptosis (36).…”

Section: Discussionmentioning

confidence: 99%

Enzymatically Inactive Procaspase 1 stabilizes the ASC Pyroptosome and Supports Pyroptosome Spreading during Cell Division

Stein

Kapplusch

Heymann

et al. 2016

Journal of Biological Chemistry

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Caspase-1 is a key player during the initiation of pro-inflammatory innate immune responses, activating pro-IL-1␤ in socalled inflammasomes. A subset of patients with recurrent febrile episodes and systemic inflammation of unknown origin harbor mutations in CASP1 encoding caspase-1. CASP1 variants result in reduced enzymatic activity of caspase-1 and impaired IL-1␤ secretion. The apparent paradox of reduced IL-1␤ secretion but systemic inflammation led to the hypothesis that CASP1 mutations may result in variable protein interaction clusters, thus activating alternative signaling pathways. To test this hypothesis, we established and characterized an in vitro system of transduced immortalized murine macrophages expressing either WT or enzymatically inactive (p.C284A) procaspase-1 fusion reporter proteins. Macrophages with variant p.C284A caspase-1 did not secrete IL-1␤ and exhibited reduced inflammatory cell death, referred to as pyroptosis. Caspase-1 and apoptosis-associated speck-like protein containing a CARD (ASC) formed cytosolic macromolecular complexes (so-called pyroptosomes) that were significantly increased in number and size in cells carrying the p.C284A caspase-1 variant compared with WT caspase-1. Furthermore, enzymatically inactive caspase-1 interacted with ASC longer and with increased intensity compared with WT caspase-1. Applying live cell imaging, we documented for the first time that pyroptosomes containing enzymatically inactive variant p.C284A caspase-1 spread during cell division. In conclusion, variant p.C284A caspase-1 stabilizes pyroptosome formation, potentially enhancing inflammation by two IL-1␤-independent mechanisms: pyroptosomes convey an enhanced inflammatory stimulus through the recruitment of additional proteins (such as RIP2, receptor interacting protein kinase 2), which is further amplified through pyroptosome and cell division.Caspase-1 is a key player during innate immune responses. It aids to control pathogens and danger signals by triggering pyroptotic cell death. Together with caspase-11 and -12 in mice and caspase-4, -5, and -12 in humans, caspase-1 belongs to the family of inflammatory caspases, which are produced as inactive forms. After the detection of pathogens or cytoplasmic danger signals, inactive caspase-1 monomers recruit to multiprotein complexes referred to as inflammasomes (1-3). In such inflammasomes, inactive procaspases are autoprocessed into caspase activation and recruitment domains (CARDs), 2 the p10 and the p20 subunits (4). Two p10 and two p20 proteins subsequently form the active caspase-1 tetramer, which then cleaves and thereby activates IL-1␤ in the so-called canonical inflammasome pathway (5, 6). Furthermore, inflammatory caspases induce pro-inflammatory cell death, referred to as pyroptosis, which involves cell swelling, cell lysis, and the release of cytoplasmic contents (7,8). Pyroptosis depends on the activation of caspase-4, -5, or -11 and caspase-1 (9 -11). Caspase-1 or -11 cleaves gasdermin D, a gasdermin domain-containing protein, at amin...

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“…The mechanisms underlying pyroptosis are unknown, but appear to initially involve the appearance of caspase-1-dependent plasma membrane pores of 1.1-2.4 nm diameter [27], followed by dissipation of ionic gradients, cell swelling, and osmotic lysis. A number of studies have highlighted the temporal association of IL-1b secretion and cell lysis [15,28,29], promoting a model in which IL-1b is passively released during cell lysis. This was most recently proposed from studies in which cytokine secretion from single cells was monitored in real-time by total internal reflection fluorescence microscopy [28,29].…”

Section: Cell Deathmentioning

confidence: 99%

Mechanisms of unconventional secretion of IL-1 family cytokines

2015

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One of the most poorly understood processes in cell biology is the peculiar ability of specific leaderless proteins to be secreted via ER/Golgi-independent mechanisms ('unconventional protein secretion'). One such leaderless protein is the major immune-activating cytokine, interleukin-1β (IL-1β). Unusual amongst cytokines, IL-1β is expressed in the cytosol as an inactive precursor protein. It requires maturation by the caspase-1 protease, which itself requires activation upon immune cell sensing of infection or cell stress. Despite 25 years of intensive research into IL-1β secretory mechanisms, how it exits the cell is still not well understood. Here we will review the various mechanisms by which macrophages have been proposed to secrete IL-1 family cytokines, and the potential involvement of caspase-1 therein. Since aberrant IL-1β production drives inherited and acquired human diseases (e.g. autoinflammatory diseases, arthritic diseases, gout, Alzheimer's disease), elucidation of the IL-1β secretory pathway may offer new therapeutic opportunities for treatment across this wide range of human conditions.

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Single-Cell Imaging of Caspase-1 Dynamics Reveals an All-or-None Inflammasome Signaling Response

Cited by 133 publications

References 38 publications

Cutting Edge: Inflammasome Activation in Primary Human Macrophages Is Dependent on Flagellin

Cutting Edge: Inflammasome Activation in Primary Human Macrophages Is Dependent on Flagellin

Enzymatically Inactive Procaspase 1 stabilizes the ASC Pyroptosome and Supports Pyroptosome Spreading during Cell Division

Mechanisms of unconventional secretion of IL-1 family cytokines

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