Hartmann Stella, Nusbaum David J, Kim Kevin, Alameh Saleem, Ho Chi-Lee C, Cruz Renae L, Levitin Anastasia, Bradley Kenneth A, Martchenko Mikhail
School of Applied Life Sciences , Keck Graduate Institute , 535 Watson Drive , Claremont , California 91711 , United States.
Department of Microbiology, Immunology and Molecular Genetics , University of California, Los Angeles , 609 Charles E. Young Drive East , Los Angeles , California 90095 , United States.
ACS Infect Dis. 2018 Dec 14;4(12):1746-1754. doi: 10.1021/acsinfecdis.8b00231. Epub 2018 Nov 5.
Inflammasomes activate caspase-1 in response to molecular signals from pathogens and other dangerous stimuli as a part of the innate immune response. A previous study discovered a small-molecule, 4-fluoro- N'-[1-(2-pyridinyl)ethylidene]benzohydrazide, which we named DN1, that reduces the cytotoxicity of anthrax lethal toxin (LT). We determined that DN1 protected cells irrespectively of LT concentration and reduced the pathogenicity of an additional bacterial exotoxin and several viruses. Using the LT cytotoxicity pathway, we show that DN1 does not prevent LT internalization and catalytic activity or caspase-1 activation. Moreover, DN1 does not affect the proteolytic activity of host cathepsin B, which facilitates the cytoplasmic entry of toxins. PubChem Bioactivities lists two G protein-coupled receptors (GPCR), type-1 angiotensin II receptor and apelin receptor, as targets of DN1. The inhibition of phosphatidylinositol 3-kinase, phospholipase C, and protein kinase B, which are downstream of GPCR signaling, synergized with DN1 in protecting cells from LT. We hypothesize that DN1-mediated antagonism of GPCRs modulates signal transduction pathways to induce a cellular state that reduces LT-induced pyroptosis downstream of caspase-1 activation. DN1 also reduced the susceptibility of Drosophila melanogaster to toxin-associated bacterial infections. Future experiments will aim to further characterize how DN1 modulates signal transduction pathways to inhibit pyroptotic cell death in LT-sensitive macrophages. DN1 represents a novel chemical probe to investigate host cellular mechanisms that mediate cell death in response to pathogenic agents.
炎性小体作为固有免疫反应的一部分，响应病原体和其他危险刺激的分子信号激活半胱天冬酶-1。先前的一项研究发现了一种小分子4-氟-N'-[1-(2-吡啶基)亚乙基]苯甲酰肼，我们将其命名为DN1，它能降低炭疽致死毒素（LT）的细胞毒性。我们确定DN1能保护细胞，与LT浓度无关，还能降低另一种细菌外毒素和几种病毒的致病性。利用LT细胞毒性途径，我们发现DN1不会阻止LT内化、催化活性或半胱天冬酶-1激活。此外，DN1不影响促进毒素进入细胞质的宿主组织蛋白酶B的蛋白水解活性。PubChem生物活性将两种G蛋白偶联受体（GPCR），即1型血管紧张素II受体和阿片肽受体列为DN1的靶点。对GPCR信号下游的磷脂酰肌醇3激酶、磷脂酶C和蛋白激酶B的抑制与DN1协同作用，保护细胞免受LT侵害。我们假设，DN1介导的GPCR拮抗作用调节信号转导途径，以诱导一种细胞状态，减少半胱天冬酶-1激活下游LT诱导的细胞焦亡。DN1还降低了黑腹果蝇对毒素相关细菌感染的易感性。未来的实验旨在进一步表征DN1如何调节信号转导途径，以抑制LT敏感巨噬细胞中的细胞焦亡性细胞死亡。DN1代表了一种新型化学探针，用于研究介导病原体引起的细胞死亡的宿主细胞机制。
