Toxoplasma gondii GRA7-Targeted ASC and PLD1 Promote Antibacterial Host Defense via PKCα.
Toxoplasma gondii GRA7-Targeted ASC and PLD1 Promote Antibacterial Host Defense via PKCα.
复制标题
弓形虫Gondii Gra7靶向ASC和PLD1通过PKCα促进抗菌宿主防御。
DOI：
10.1371/journal.ppat.1006126
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发表时间：
2017-01
期刊：
影响因子：
6.7
通讯作者：
Yang CS
中科院分区：
文献类型：
--
作者：
Koh HJ;Kim YR;Kim JS;Yun JS;Jang K;Yang CS
结核病是一个全球性的健康问题，至少有三分之一的世界人口感染结核分枝杆菌（MTB）。结核分枝杆菌是一种成功的病原体，通过抑制炎症和阻止吞噬-溶酶体融合来增强其自身的细胞内存活。我们以前证明弓形虫（T。弓形虫）致密颗粒抗原（GRA）7通过髓样分化初级应答基因88与TNF受体相关因子6相互作用，使得巨噬细胞中的先天免疫应答成为可能。为了扩展这些研究，我们发现GRA 7与参与抗微生物宿主防御机制的宿主蛋白相互作用，作为结核病的治疗策略。在这里，我们发现蛋白激酶C（PKC）α介导的T。弓形虫GRA 7-I（Ser 52）调控GRA 7与结核病相关斑点样蛋白PYD结构域的相互作用，该蛋白含有羧基末端CARD，能够寡聚化，并且炎性小体激活可以导致针对MTB的抗菌防御。此外，GRA 7-III与磷脂酶D1的PX结构域相互作用，通过PKCα以GRA 7-III（Ser 135）磷酸化依赖的方式促进其酶活性、吞噬溶酶体成熟和随后的抗菌活性。总之，这些结果强调了以前未被认识到的作用，GRA 7在调节抗微生物宿主防御机制，在分枝杆菌感染。我们以前证明弓形虫（T。弓形虫致密颗粒抗原（GRA）7通过MyD 88与TRAF 6相互作用，使巨噬细胞中的先天性免疫应答和有效的抗弓形虫保护作用成为可能。体内弓形虫感染。然而，它的确切作用以及它如何调节宿主先天免疫反应尚未得到充分解释。在此，我们发现PKCα介导的GRA 7磷酸化对于GRA 7与ASC或PLD 1之间的相互作用是必需的，这可以促进对结核分枝杆菌（MTB）的抗菌防御。值得注意的是，在体外和体内，PKCα特异性磷酸化GRA 7的Ser 52和Ser 135，表明GRA 7是PKCα的底物。GRA 7的N-末端（GRA 7-I）足以与ASC的PYD结构域相互作用，其能够进行ASC寡聚化和炎性小体活化。此外，GRA 7-III与PLD 1的PX结构域相互作用，以GRA 7磷酸化依赖的方式促进其酶活性、吞噬溶酶体成熟和随后的抗微生物活性。有趣的是，GRA 7中的磷酸化模拟突变克服了对PKCα的需要。总的来说，这些结果为GRA 7如何以PKCα依赖性方式促进ASC和PLD 1活化作为抗微生物宿主防御机制提供了新的见解。
Tuberculosis is a global health problem and at least one-third of the world’s population is infected with Mycobacterium tuberculosis (MTB). MTB is a successful pathogen that enhances its own intracellular survival by inhibiting inflammation and arresting phago-lysosomal fusion. We previously demonstrated that Toxoplasma gondii (T. gondii) dense granule antigen (GRA) 7 interacts with TNF receptor-associated factor 6 via Myeloid differentiation primary response gene 88, enabling innate immune responses in macrophages. To extend these studies, we found that GRA7 interacts with host proteins involved in antimicrobial host defense mechanisms as a therapeutic strategy for tuberculosis. Here, we show that protein kinase C (PKC)α-mediated phosphorylation of T. gondii GRA7-I (Ser52) regulates the interaction of GRA7 with PYD domain of apoptosis-associated speck-like protein containing a carboxy-terminal CARD, which is capable of oligomerization and inflammasome activation can lead to antimicrobial defense against MTB. Furthermore, GRA7-III interacted with the PX domain of phospholipase D1, facilitating its enzyme activity, phago-lysosomal maturation, and subsequent antimicrobial activity in a GRA7-III (Ser135) phosphorylation-dependent manner via PKCα. Taken together, these results underscore a previously unrecognized role of GRA7 in modulating antimicrobial host defense mechanism during mycobacterial infection. We previously demonstrated that Toxoplasma gondii (T. gondii) dense granule antigen (GRA) 7 interacts with TRAF6 via MyD88, enabling innate immune responses in macrophages and effective protection against T. gondii infection in vivo. However, its exact role and how it regulates host innate immune responses have not been fully explained. Herein, we show that PKCα-mediated phosphorylation of GRA7 is essential for the interaction between GRA7 and ASC or PLD1, which can promote antimicrobial defense against Mycobacterium tuberculosis (MTB). Notably, PKCα specifically phosphorylated Ser52 and Ser135 of GRA7 in vitro and in vivo, indicating that GRA7 is a substrate of PKCα. The N-terminal of GRA7 (GRA7-I) was sufficient for interaction with the PYD domain of ASC, which is capable of ASC oligomerization and inflammasome activation. Furthermore, GRA7-III interacted with the PX domain of PLD1, facilitating its enzyme activity, phago-lysosomal maturation, and subsequent antimicrobial activity in a GRA7 phosphorylation-dependent manner. Interestingly, phosphomimetic mutation in GRA7 overcame the need for PKCα. Collectively, these results provide novel insight into how GRA7 can promote ASC and PLD1 activation in a PKCα-dependent manner as an antimicrobial host defense mechanism.