Comunicación

ACTIVATION AND REGULATION OF THE NLRP3 INFLAMMASOME WITH MUTATIONS LINKED TO CRYOPYRIN-ASSOCIATED PERIODIC SYNDROMES

Autores:

MARÍA CRISTINA MOLINA LÓPEZ1, DIEGO ANGOSTO BAZARRA1, ANA TAPIA ABELLÁN1, LAURA HURTADO NAVARRO1, PABLO PELEGRIN VIVANCOS1

Afiliaciones:

(1) CIRUGÍA DIGESTIVA, ENDOCRINA Y TRASPLANTE DE ÓRGANOS ABDOMINALES, IMIB-Arrixaca, España

Comunicación:

Antecedentes:

Cryopyrin-associated periodic syndrome (CAPS) is a hereditary autoinflammatory disorder that results from the gain-of-function mutations in the NLRP3 gene resulting in the autoactivation of the NLRP3 inflammasome and an overproduction of the proinflammatory cytokines IL-1β and IL-18. In wild-type, different pathogen and host-related signals are required to activate the NLRP3 inflammasome, which is formed by NLPR3, the adaptor protein ASC and the effector enzyme pro-caspase-1. Once the inflammasome is formed, pro-caspase-1 is activated and cleaves pro-IL-1β and pro-IL-18 to their active forms and cleaves gasdermin D which then its N-terminal fragment form pores in the plasma membrane, through where IL-1β, IL-18 and intracellular content are released. Gasdermin D pores then drives a specific type of cell death called pyroptosis. However, the activation and regulation mechanism of mutant NLRP3 is still poorly known. Questions such as how the activity of mutated NLRP3 is regulated in sterile conditions in patients; whether MCC950 (a specific inhibitor of NLRP3) is able to inhibit this activation; what molecules besides IL-1β and IL-18 are implicated downstream the activation pathway, and how NLRP3 mutant is regulated at post-translational level are still unknown.

Métodos:

Nlrp3-deficient immortalized mouse macrophages with a Tet-on inducible system were used to express several NLRP3 variants with mutations present in CAPS patients and compared to macrophages transduced with empty vector or with wild-type NLRP3.CAPS patient blood was used to study IL-1β production and formation of ASC oligomers as readouts of NLRP3 activation. Cells were treated with LPS, Pam3CSK4, IL-6, S100A9, IL-1α, TNF-α, palmitate, ATP, or uric acid crystals at different concentrations and times in the absence or presence of MCC950 or with different deubiquitinase inhibitors.

Resultados:

Contrasting with the expression of wild-type NLRP3, the sole expression of mutant NLRP3 results in a constitutive activation of the inflammasome with the activation of caspase-1 and gasdermin D cleavage, followed by the release of several cellular proteins as IL-18, P2X7 or LDH. IL-1β release was only observed after LPS, Pam3CSK4, IL-6, palmitate and S100A9 treatment, compounds that activate NF-kB and the expression of Il1b gene.MCC950 was able to block the release of IL-1β, IL-18, HMGB1, IL-1α, P2X7, cystatin B, annexin 1 but not TNF-a, confirming NLRP3 activation. Deubiquitinase inhibitors were able to reduce NLRP3 activation and IL-1β release when mutant NLRP3 was expressed, suggesting that ubiquitination of mutant NLRP3 restricts its activity.

Conclusiones:

Gain-of-function NLRP3 mutations present a basal activation with respect to wild-type NLPR3, and its activation tightly depends on its expression levels, but could be further increased by different endogenous host-derived signals, such as IL-6, palmitate or S100A9 proteins. Additionally, mutated auto-active NLRP3 is post-translational regulated by ubiquitination, that impairs auto-activation. This study shed new light into the origins of non-infectious inflammatory flares in CAPS patients.


Dirección

Campus de Ciencias de la Salud
Carretera Buenavista s/n, 30120 El Palmar
Murcia, España

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