Toxoplasma gondii-induced activation of EGFR prevents autophagy protein-mediated killing of the parasite.
Toxoplasma gondii-induced activation of EGFR prevents autophagy protein-mediated killing of the parasite.
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DOI：
10.1371/journal.ppat.1003809
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发表时间：
2013
期刊：
影响因子：
6.7
通讯作者：
Subauste CS
中科院分区：
文献类型：
--
作者：
Muniz-Feliciano L;Van Grol J;Portillo JA;Liew L;Liu B;Carlin CR;Carruthers VB;Matthews S;Subauste CS
刚地弓形虫存在于细胞内隔室（寄生虫空泡），排除了内体-溶酶体募集所需的跨膜分子。因此，寄生虫通过避免溶酶体降解而存活。然而，自噬可以将寄生虫空泡重新路由到溶酶体并导致寄生虫杀死。这就提出了T.弓形虫可能采用一种策略来防止自噬靶向，以维持空泡的非融合性质。我们报告T.在内皮细胞、视网膜色素上皮细胞和小胶质细胞中，弓形虫激活EGFR。EGFR或其下游分子Akt的阻断通过依赖于自噬蛋白Atg 7和Beclin 1的LC 3+结构、空泡-溶酶体融合、溶酶体降解和寄生虫杀伤引起寄生虫的靶向。阻断GPCR或抑制金属蛋白酶并不能阻止EGFR-Akt的激活。T.含有EGF结构域的弓形虫微线蛋白（MIC）（EGF-MIC; MIC 3和MIC 6）似乎促进EGFR活化。EGF-MIC缺陷的寄生虫（MIC 1 ko，MIC 1缺陷和MIC 6分泌; MIC 3 ko，MIC 3缺陷;和MIC 1 -3 ko，MIC 1、MIC 3缺陷和MIC 6分泌）导致EGFR-Akt活化受损，重组EGF-MIC（MIC 3和MIC 6）导致EGFR-Akt活化。在用自噬刺激剂（CD 154，雷帕霉素）处理的细胞中，EGFR信号传导抑制寄生虫周围的LC 3积累。此外，增加的LC 3积累和寄生虫杀死中注意到CD 154激活的细胞感染MIC 1 -3 ko寄生虫。最后，重组MIC 3和MIC 6抑制由CD 154触发的寄生虫杀伤，特别是针对MIC 1 -3 ko寄生虫。因此，我们的研究结果确定EGFR激活作为T。维持寄生虫空泡的非融合性质，并表明EGF-MIC在影响宿主细胞中的信号传导以促进寄生虫存活方面具有新的作用。 刚地弓形虫存在于寄生虫的空泡中，该空泡排除了内体和溶酶体募集所需的跨膜蛋白，因此不遵循经典的溶酶体降解路径。然而，当自噬（一种溶酶体降解途径）通过免疫系统或溶酶体上调时，液泡的非融合性质可以恢复。维持液泡的非融合性是寄生虫生存的关键。因此，除了通过经典的溶酶体途径防止降解外，T.弓形虫还可以部署策略以防止自噬的组成性水平靶向病原体并引起其溶酶体降解。我们报告T.弓形虫通过在宿主细胞中引起EGFR活化来完成这一任务。在未经历自噬的免疫或药理学上调的细胞中，EGFR的阻断导致寄生虫被表达自噬蛋白LC 3的结构包裹，空泡-溶酶体融合和寄生虫的自噬蛋白依赖性杀伤。此外，EGFR信号传导还损害了用自噬刺激剂处理的细胞中LC 3+结构对寄生虫的靶向。研究与T。缺乏EGF结构域的弓形虫-微线蛋白（EGF-MIC）和重组EGF-MIC支持这些寄生虫粘附素有助于EGFR活化的概念。
Toxoplasma gondii resides in an intracellular compartment (parasitophorous vacuole) that excludes transmembrane molecules required for endosome - lysosome recruitment. Thus, the parasite survives by avoiding lysosomal degradation. However, autophagy can re-route the parasitophorous vacuole to the lysosomes and cause parasite killing. This raises the possibility that T. gondii may deploy a strategy to prevent autophagic targeting to maintain the non-fusogenic nature of the vacuole. We report that T. gondii activated EGFR in endothelial cells, retinal pigment epithelial cells and microglia. Blockade of EGFR or its downstream molecule, Akt, caused targeting of the parasite by LC3+ structures, vacuole-lysosomal fusion, lysosomal degradation and killing of the parasite that were dependent on the autophagy proteins Atg7 and Beclin 1. Disassembly of GPCR or inhibition of metalloproteinases did not prevent EGFR-Akt activation. T. gondii micronemal proteins (MICs) containing EGF domains (EGF-MICs; MIC3 and MIC6) appeared to promote EGFR activation. Parasites defective in EGF-MICs (MIC1 ko, deficient in MIC1 and secretion of MIC6; MIC3 ko, deficient in MIC3; and MIC1-3 ko, deficient in MIC1, MIC3 and secretion of MIC6) caused impaired EGFR-Akt activation and recombinant EGF-MICs (MIC3 and MIC6) caused EGFR-Akt activation. In cells treated with autophagy stimulators (CD154, rapamycin) EGFR signaling inhibited LC3 accumulation around the parasite. Moreover, increased LC3 accumulation and parasite killing were noted in CD154-activated cells infected with MIC1-3 ko parasites. Finally, recombinant MIC3 and MIC6 inhibited parasite killing triggered by CD154 particularly against MIC1-3 ko parasites. Thus, our findings identified EGFR activation as a strategy used by T. gondii to maintain the non-fusogenic nature of the parasitophorous vacuole and suggest that EGF-MICs have a novel role in affecting signaling in host cells to promote parasite survival. Toxoplasma gondii resides in a parasitophorous vacuole that excludes transmembrane proteins required for recruitment of endosomes and lysosomes and thus, does not follow the path of classical lysosomal degradation. However, the non-fusogenic nature of the vacuole can be reverted when autophagy, a pathway to lysosomal degradation, is upregulated through the immune system or pharmacologically. Maintenance of the non-fusogenic nature of the vacuole is central to parasite survival. Thus, in addition to preventing degradation through a classical lysosomal pathway, T. gondii may also deploy strategies to prevent constitutive levels of autophagy from targeting the pathogen and causing its lysosomal degradation. We report that T. gondii accomplishes this task by causing EGFR activation in host cells. In cells that were not subjected to immune or pharmacologic upregulation of autophagy, blockade of EGFR resulted in parasite encasing by structures that expressed the autophagy protein LC3, vacuole-lysosomal fusion and autophagy protein-dependent killing of the parasite. Moreover, EGFR signaling also impaired targeting of the parasite by LC3+ structures in cells treated with stimulators of autophagy. Studies with T. gondii deficient in EGF domain containing-micronemal proteins (EGF-MICs) and recombinant EGF-MICs support the concept that these parasite adhesins contribute to EGFR activation.
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DOI：
10.1126/science.1227026
发表时间：
2012-11-23
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Choy A;Dancourt J;Mugo B;O'Connor TJ;Isberg RR;Melia TJ;Roy CR
通讯作者：
Roy CR
影响因子：
32.4
作者：
Blanchet FP;Moris A;Nikolic DS;Lehmann M;Cardinaud S;Stalder R;Garcia E;Dinkins C;Leuba F;Wu L;Schwartz O;Deretic V;Piguet V
通讯作者：
Piguet V
影响因子：
64.5
作者：
Coppens, I;Dunn, JD;Joiner, KA
通讯作者：
Joiner, KA
微生物蛋白在弓形虫贡献毒力中的协同作用。
影响因子：
15.3
作者：
Céréde, O;Dubremetz, JF;Soête, M;Deslée, D;Vial, H;Bout, D;Lebrun, M
通讯作者：
Lebrun, M
影响因子：
56.9
作者：
JOINER, KA;FUHRMAN, SA;MELLMAN, I
通讯作者：
MELLMAN, I