Stimulation of ectopically expressed muscarinic receptors induces IFN-γ but suppresses IL-2 production by inhibiting activation of pAKT pathways in primary T cells.
Stimulation of ectopically expressed muscarinic receptors induces IFN-γ but suppresses IL-2 production by inhibiting activation of pAKT pathways in primary T cells.
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DOI：
10.1073/pnas.2300987120
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发表时间：
2023-06-20
影响因子：
11.1
通讯作者：
Weiss, Arthur
中科院分区：
文献类型：
--
作者：
Nguyen, Trang T. T.;Lu, Wen;Zhu, Wandi S.;Ansel, K. Mark;Erh Liang, Hong-;Weiss, Arthur
研究G蛋白偶联的毒蕈碱受体的功能，这是蛋白酪氨酸激酶的独立性，可能会激发新的癌症治疗绕过经典的TCR信号通路。如果PLCβ1共表达，刺激异源表达的毒蕈碱受体（M1和合成的hM 3Dq）诱导预活化T细胞中的钙反应和ERK磷酸化。出乎意料的是，尽管与PLCβ1偶联的hM 3Dq的刺激诱导高IFN-γ、CD 69和CD 25表达，但令人惊讶的是，它不诱导高IL-2表达。hM 3Dq的刺激降低了IL-2 mRNA的稳定性，这与对IL-2 mRNA稳定性的影响相关。对IL-2 mRNA的选择性作用可能归因于pAKT下游途径功能的降低，表明pAKT途径对IL-2产生至关重要。T细胞抗原受体刺激诱导下游信号分子和磷脂酰肌醇、Ras、MAPK和PI 3激酶途径的酪氨酸磷酸化，导致T细胞活化。以前，我们报道了G蛋白偶联的人毒蕈碱受体可以绕过酪氨酸激酶激活磷脂酰肌醇途径，诱导Jurkat白血病T细胞产生白细胞介素-2。在这里，我们证明，刺激G蛋白偶联的毒蕈碱受体（M1和合成的hM 3Dq）可以激活原代小鼠T细胞，如果PLCβ1共表达。静息的外周血hM 3Dq +PLCβ1（hM 3Dq/β1）T细胞对hM 3Dq激动剂氯氮平无反应，除非它们被TCR和CD 28刺激预激活，从而增加hM 3Dq和PLCβ1的表达。这允许大量的钙和磷酸化ERK对氯氮平的反应。氯氮平处理诱导高IFN-γ、CD 69和CD 25表达，但令人惊讶地在hM 3Dq/β1 T细胞中不诱导大量IL-2。重要的是，两种毒蕈碱受体加上TCR的共刺激甚至导致IL-2表达减少，表明毒蕈碱受体共刺激的选择性抑制作用。刺激毒蕈碱受体可诱导NFAT和NFκB的强烈核转位并激活AP-1。而hM 3Dq的刺激导致IL-2 mRNA稳定性降低，这与IL-2 3′UTR活性的影响有关。有趣的是，hM 3Dq的刺激导致pAKT及其下游途径减少。这可能解释了对hM 3Dq/β 1 T细胞中IL-2产生的抑制作用。此外，PI 3 K的抑制剂减少了TCR刺激的hM 3Dq/β1 CD 4 T细胞中的IL-2产生，表明激活pAKT途径对于T细胞中的IL-2产生至关重要。
Studying the functions of G-protein-coupled muscarinic receptors, which are protein tyrosine kinase independent, might inspire new cancer therapies by bypassing classical TCR signaling pathways. Stimulating heterologously expressed muscarinic receptors (M1 and the synthetic hM3Dq) induced calcium responses and phosphorylation of ERK in preactivated T cells if PLCβ1 was coexpressed. Unexpectedly, whereas stimulation of hM3Dq that couples to PLCβ1 induced high IFN-γ, CD69, and CD25 expression, surprisingly, it did not induce high IL-2 expression. Stimulation of hM3Dq reduced IL-2 mRNA stability which correlated with an effect on the IL-2 mRNA stability. The selective effect on IL-2 mRNA may be attributable to reduced pAKT downstream pathway function, suggesting that the pAKT pathway is critical for IL-2 production. T cell antigen receptor stimulation induces tyrosine phosphorylation of downstream signaling molecules and the phosphatidylinositol, Ras, MAPK, and PI3 kinase pathways, leading to T cell activation. Previously, we reported that the G-protein-coupled human muscarinic receptor could bypass tyrosine kinases to activate the phosphatidylinositol pathway and induce interleukin-2 production in Jurkat leukemic T cells. Here, we demonstrate that stimulating G-protein-coupled muscarinic receptors (M1 and synthetic hM3Dq) can activate primary mouse T cells if PLCβ1 is coexpressed. Resting peripheral hM3Dq+PLCβ1 (hM3Dq/β1) T cells did not respond to clozapine, an hM3Dq agonist, unless they were preactivated by TCR and CD28 stimulation which increased hM3Dq and PLCβ1 expression. This permitted large calcium and phosphorylated ERK responses to clozapine. Clozapine treatment induced high IFN-γ, CD69, and CD25 expression, but surprisingly did not induce substantial IL-2 in hM3Dq/β1 T cells. Importantly, costimulation of both muscarinic receptors plus the TCR even led to reduced IL-2 expression, suggesting a selective inhibitory effect of muscarinic receptor costimulation. Stimulation of muscarinic receptors induced strong nuclear translocation of NFAT and NFκB and activated AP-1. However, stimulation of hM3Dq led to reduced IL-2 mRNA stability which correlated with an effect on the IL-2 3′UTR activity. Interestingly, stimulation of hM3Dq resulted in reduced pAKT and its downstream pathway. This may explain the inhibitory impact on IL-2 production in hM3Dq/β1T cells. Moreover, an inhibitor of PI3K reduced IL-2 production in TCR-stimulated hM3Dq/β1 CD4 T cells, suggesting that activating the pAKT pathway is critical for IL-2 production in T cells.
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DOI：
10.1038/nrd4295
发表时间：
2014-07
期刊：
Nature reviews. Drug discovery
影响因子：
--
作者：
Kruse AC;Kobilka BK;Gautam D;Sexton PM;Christopoulos A;Wess J
通讯作者：
Wess J
DOI：
10.4049/jimmunol.1100820
发表时间：
2011-08-01
期刊：
Journal of immunology (Baltimore, Md. : 1950)
影响因子：
--
作者：
Kremer KN;Clift IC;Miamen AG;Bamidele AO;Qian NX;Humphreys TD;Hedin KE
通讯作者：
Hedin KE
关闭 AKT：PHLPP 作为药物靶点。
DOI：
10.1146/annurev-pharmtox-011112-140338
发表时间：
2014
影响因子：
12.5
作者：
Newton AC;Trotman LC
通讯作者：
Trotman LC
影响因子：
64.5
作者：
Manning BD;Toker A
通讯作者：
Toker A
影响因子：
4.4
作者：
Pei，Yuan;Zhu，Ping;Yu，Long
通讯作者：
Yu，Long