Genetic targeting of Card19 is linked to disrupted NINJ1 expression, impaired cell lysis, and increased susceptibility to Yersinia infection.
Genetic targeting of Card19 is linked to disrupted NINJ1 expression, impaired cell lysis, and increased susceptibility to Yersinia infection.
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DOI：
10.1371/journal.ppat.1009967
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发表时间：
2021-10
期刊：
影响因子：
6.7
通讯作者：
Brodsky IE
中科院分区：
文献类型：
--
作者：
Bjanes E;Sillas RG;Matsuda R;Demarco B;Fettrelet T;DeLaney AA;Kornfeld OS;Lee BL;Rodríguez López EM;Grubaugh D;Wynosky-Dolfi MA;Philip NH;Krespan E;Tovar D;Joannas L;Beiting DP;Henao-Mejia J;Schaefer BC;Chen KW;Broz P;Brodsky IE
细胞死亡在炎症反应中起关键作用。在焦亡过程中，炎性半胱天冬酶裂解气凝胶蛋白D (GSDMD)，释放一个n端片段，产生质膜孔，介导细胞裂解和IL-1细胞因子释放。在某些细胞类型中或在特定刺激下，caspase激活后的终端细胞裂解和IL-1β释放可以解耦，这种状态称为超激活。然而，调控GSDMD裂解下游末端细胞裂解的因素和机制仍然知之甚少。在研究耶尔森菌感染过程中焦亡调控的过程中，我们鉴定了一株card19缺陷小鼠（Card19lxcn），其巨噬细胞免受细胞裂解的保护，细胞凋亡和焦亡减少，但caspase激活、IL-1分泌和GSDMD切割水平均为野生型。出乎意料的是，线粒体中含有CARD19的蛋白CARD19并不是直接导致这一现象的原因，因为一个独立产生的CRISPR/Cas9敲除CARD19的小鼠系（Card19Null）在巨噬细胞裂解中没有表现出缺陷。值得注意的是，Card19位于13号染色体上，紧挨着最近被发现调控GSDMD激活下游细胞裂解的Ninj1。RNA-seq和western blotting结果显示，Card19lxcn永活BMDMs显著降低了NINJ1的表达，并且Card19lxcn永活BMDMs中NINJ1的重组恢复了它们在caspase依赖性细胞死亡刺激下进行细胞分解的能力。Card19lxcn小鼠对耶尔森菌感染的易感性增加，而独立生成的Card19Null小鼠则没有，这表明细胞裂解本身在抵抗细菌感染中起关键作用，Card19lxcn小鼠的感染易感性增加可归因于NINJ1的缺失。我们的研究发现，Card19的基因靶向对相邻基因Ninj1产生脱靶效应，破坏巨噬细胞在气皮蛋白裂解下游进行质膜破裂的能力，并影响宿主生存和耶尔森菌感染期间的细菌控制。程序性细胞死亡是调节组织稳态和宿主防御感染的关键。焦亡是一种程序性细胞死亡的炎症形式，伴随着细胞裂解和炎性细胞因子的释放。细胞裂解是由特定细胞内孔形成蛋白的激活触发的，但细胞裂解的调控机制尚不清楚。Card19在13号染色体上的遗传靶向导致相邻基因Ninj1的表达减少，该基因最近被发现在响应细胞死亡诱导刺激时调节末端裂解事件。一致地，来自card19缺陷小鼠的巨噬细胞对多种炎症刺激的多种形式的细胞死亡具有抗性，包括典型和非典型炎性体激活，以及细胞外源性凋亡的触发。值得注意的是，card19缺陷小鼠更容易感染耶尔森氏菌，这表明细胞裂解有助于控制细菌感染。我们的数据为研究终端细胞裂解对细菌感染控制的影响提供了新的见解，并强调了在气真皮蛋白裂解下游调节裂解细胞死亡的其他因素的作用。
Cell death plays a critical role in inflammatory responses. During pyroptosis, inflammatory caspases cleave Gasdermin D (GSDMD) to release an N-terminal fragment that generates plasma membrane pores that mediate cell lysis and IL-1 cytokine release. Terminal cell lysis and IL-1β release following caspase activation can be uncoupled in certain cell types or in response to particular stimuli, a state termed hyperactivation. However, the factors and mechanisms that regulate terminal cell lysis downstream of GSDMD cleavage remain poorly understood. In the course of studies to define regulation of pyroptosis during Yersinia infection, we identified a line of Card19-deficient mice (Card19lxcn) whose macrophages were protected from cell lysis and showed reduced apoptosis and pyroptosis, yet had wild-type levels of caspase activation, IL-1 secretion, and GSDMD cleavage. Unexpectedly, CARD19, a mitochondrial CARD-containing protein, was not directly responsible for this, as an independently-generated CRISPR/Cas9 Card19 knockout mouse line (Card19Null) showed no defect in macrophage cell lysis. Notably, Card19 is located on chromosome 13, immediately adjacent to Ninj1, which was recently found to regulate cell lysis downstream of GSDMD activation. RNA-seq and western blotting revealed that Card19lxcn BMDMs have significantly reduced NINJ1 expression, and reconstitution of Ninj1 in Card19lxcn immortalized BMDMs restored their ability to undergo cell lysis in response to caspase-dependent cell death stimuli. Card19lxcn mice exhibited increased susceptibility to Yersinia infection, whereas independently-generated Card19Null mice did not, demonstrating that cell lysis itself plays a key role in protection against bacterial infection, and that the increased infection susceptibility of Card19lxcn mice is attributable to loss of NINJ1. Our findings identify genetic targeting of Card19 being responsible for off-target effects on the adjacent gene Ninj1, disrupting the ability of macrophages to undergo plasma membrane rupture downstream of gasdermin cleavage and impacting host survival and bacterial control during Yersinia infection. Programmed cell death is critical for regulating tissue homeostasis and host defense against infection. Pyroptosis is an inflammatory form of programmed cell death that couples cell lysis with release of inflammatory cytokines. Cell lysis is triggered by activation of particular intracellular pore forming proteins, but how regulation of cell lysis occurs is not well understood. Genetic targeting of Card19 on chromosome 13 resulted in decreased expression of the adjacent gene, Ninj1, which was recently found to regulate terminal lysis events in response to cell death-inducing stimuli. Consistently, macrophages from Card19-deficient mice were resistant to multiple forms of cell death in response to a variety of inflammatory stimuli, including canonical and non-canonical inflammasome activation, as well as triggers of cell-extrinsic apoptosis. Notably, Card19-deficient mice were more susceptible to Yersinia infection, indicating that cell lysis contributes to control of bacterial infections. Our data provide new insight into the impact of terminal cell lysis on control of bacterial infection and highlight the role of additional factors that regulate lytic cell death downstream of gasdermin cleavage.
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形成孔的蛋白质加油D可以调节白细胞介素-1的巨噬细胞分泌。
影响因子：
32.4
作者：
Evavold CL;Ruan J;Tan Y;Xia S;Wu H;Kagan JC
通讯作者：
Kagan JC
DOI：
10.1073/pnas.1607769113
发表时间：
2016-07-12
影响因子：
11.1
作者：
Aglietti, Robin A.;Estevez, Alberto;Dueber, Erin C.
通讯作者：
Dueber, Erin C.
影响因子：
12.4
作者：
Conos, S. A.;Lawlor, K. E.;Lindqvist, L. M.
通讯作者：
Lindqvist, L. M.
影响因子：
30.5
作者：
Franchi, Luigi;Amer, Amal;Nunez, Gabriel
通讯作者：
Nunez, Gabriel
影响因子：
14.8
作者：
Degterev, Alexei;Hitomi, Junichi;Yuan, Junying
通讯作者：
Yuan, Junying