Leishmania donovani induced Unfolded Protein Response delays host cell apoptosis in PERK dependent manner.
Leishmania donovani induced Unfolded Protein Response delays host cell apoptosis in PERK dependent manner.
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DOI：
10.1371/journal.pntd.0006646
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发表时间：
2018-07
影响因子：
3.8
通讯作者：
Das P
中科院分区：
文献类型：
--
作者：
Abhishek K;Das S;Kumar A;Kumar A;Kumar V;Saini S;Mandal A;Verma S;Kumar M;Das P
内质网（ER）应激产生的未折叠应激反应（UPR）是细胞在不利条件下保护细胞的一种基本生存机制。利什曼原虫以各种方式调节宿主巨噬细胞以确保其存活。在宿主的延迟凋亡诱导中调节PI 3 K-Akt途径;使寄生虫能够稳定感染以进一步繁殖。将感染的RAW巨噬细胞暴露于喜树碱或thagsigargin，并使用磷酸特异性抗体通过蛋白质印迹法测定PERK、Akt、BAD和Cyt-C的磷酸化状态。使用真实的时间PCR测定cIAP 1和2、ATF 4、CHOP、ATF 3、HO-1和sXBP 1在转录水平上的表达。对于抑制研究，在感染前用PERK抑制剂GSK 2606414预处理RAW巨噬细胞。我们在RAW巨噬细胞中的研究表明，针对杜氏乳杆菌感染的宿主UPR的诱导激活了Akt介导的通路，其延迟了宿主的凋亡诱导。此外，利什曼原虫感染导致宿主PERK酶的磷酸化和激活以及凋亡抑制基因家族（cIAP）mRNA的基因转录增加。在我们的抑制研究中，我们发现在凋亡诱导剂下抑制感染诱导的PERK磷酸化降低了Akt磷酸化，并且未能激活参与抗凋亡保护的进一步下游分子。此外，在氧化暴露下抑制PERK磷酸化导致一氧化氮产生增加。同时，PERK磷酸化后cIAP mRNA的转录减少导致宿主细胞凋亡，从而降低感染率。总的来说，该研究的结果表明，利什曼原虫调节宿主UPR和PERK磷酸化延迟宿主巨噬细胞中的凋亡诱导，因此支持寄生虫在感染早期阶段的入侵。内脏利什曼病（又称黑热病）是由杜氏利什曼原虫（Leishmaniadonovani）引起的印度次大陆的一种严重的热带寄生虫病。宿主在延迟凋亡诱导方面的调节是有利于疾病建立的方面之一;然而，其机制尚不清楚。本研究旨在探讨杜氏乳杆菌感染诱导的UPR与细胞凋亡延迟发生之间的关系。我们发现杜氏乳杆菌感染使宿主体内PERK和Akt分子磷酸化，并伴有细胞凋亡延迟。同时，在感染的巨噬细胞中，细胞IAP（cIAP 1和2）基因的水平也上调。为了评估PERK在宿主延迟凋亡中的参与，我们在暴露于凋亡诱导剂的情况下抑制PERK的磷酸化。我们发现，PERK抑制降低Akt磷酸化，并未能激活其他相关的下游分子参与延迟宿主凋亡。此外，观察到cIAP水平显著降低。在氧化暴露下，PERK磷酸化的抑制削弱了受感染的RAW细胞维持氧化还原稳态的能力，导致更高的一氧化氮产生。总之，杜氏乳杆菌感染以PERK依赖性方式调节宿主细胞凋亡并有利于感染。
Endoplasmic reticulum (ER) stress generated unfolded stress response (UPR) is a basic survival mechanism which protects cell under unfavourable conditions. Leishmania parasite modulates host macrophages in various ways to ensure its survival. Modulation of PI3K-Akt pathway in delayed apoptotic induction of host; enables parasite to stabilize the infection for further propagation. Infected RAW macrophages were exposed to campothecin or thagsigargin and phosphorylation status of PERK, Akt, BAD and Cyt-C was determined through western blotting using phospho specific antibody. Expression at transcriptional level for cIAP1 &2, ATF4, CHOP, ATF3, HO-1 and sXBP1 was determined using real time PCR. For inhibition studies, RAW macrophages were pre-treated with PERK inhibitor GSK2606414 before infection. Our studies in RAW macrophages showed that induction of host UPR against L.donovani infection activates Akt mediated pathway which delays apoptotic induction of the host. Moreover, Leishmania infection results in phosphorylation and activation of host PERK enzyme and increased transcription of genes of inhibitor of apoptosis gene family (cIAP) mRNA. In our inhibition studies, we found that inhibition of infection induced PERK phosphorylation under apoptotic inducers reduces the Akt phosphorylation and fails to activate further downstream molecules involved in protection against apoptosis. Also, inhibition of PERK phosphorylation under oxidative exposure leads to increased Nitric Oxide production. Simultaneously, decreased transcription of cIAP mRNA upon PERK phosphorylation fates the host cell towards apoptosis hence decreased infection rate. Overall the findings from the study suggests that Leishmania modulated host UPR and PERK phosphorylation delays apoptotic induction in host macrophage, hence supports parasite invasion at early stages of infection. Visceral Leishmaniasis or Kala-azar is one of the severe tropical neglected parasitic diseases caused by Leishmania donovani in Indian subcontinent. Modulation of host in terms of delayed apoptotic induction is one of the aspects which favours disease establishment; however the mechanism is not clearly understood yet. In the present study, we tried to explore the connection between L.donovani infection induced UPR in host with delayed onset of apoptosis. We found that L.donovani infection phosphorylates the PERK and Akt molecule in host along with delayed apoptosis. Simultaneously, the levels of cellular IAP (cIAP1 & 2) genes were also up-regulated in infected macrophages. To assess the involvement of PERK in delayed apoptosis of host, we inhibited the phosphorylation of PERK under the exposure to apoptotic inducers. We found that PERK inhibition decreased the Akt phosphorylation and fails to activate other associated downstream molecules involved in delayed apoptosis of host. Also, a significant reduction in cIAP levels was observed. Under oxidative exposure, inhibition of PERK phosphorylation debilitates infected RAW cell’s ability to maintain redox homeostasis leading to higher nitric oxide production. Altogether, L.donovani infection modulates host apoptosis in a PERK dependent manner and favours infection.
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影响因子：
7.4
作者：
Circu, Magdalena L.;Aw, Tak Yee
通讯作者：
Aw, Tak Yee
DOI：
10.1016/0169-4758(96)10067-3
发表时间：
1996-11-01
期刊：
PARASITOLOGY TODAY
影响因子：
--
作者：
Brittingham, A;Mosser, DM
通讯作者：
Mosser, DM
影响因子：
16
作者：
Harding, HP;Zhang, YH;Ron, D
通讯作者：
Ron, D
影响因子：
3.4
作者：
Dermine, JF;Scianimanico, S;Desjardins, M
通讯作者：
Desjardins, M
影响因子：
3.7
作者：
Galluzzi L;Diotallevi A;De Santi M;Ceccarelli M;Vitale F;Brandi G;Magnani M
通讯作者：
Magnani M