Intracellular arginine-dependent translation sensor reveals the dynamics of arginine starvation response and resistance in ASS1-negative cells.
Intracellular arginine-dependent translation sensor reveals the dynamics of arginine starvation response and resistance in ASS1-negative cells.
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DOI：
10.1186/s40170-021-00238-9
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发表时间：
2021-01-21
影响因子：
5.9
通讯作者：
Van Tine BA
中科院分区：
文献类型：
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作者：
Rogers LC;Zhou J;Baker A;Schutt CR;Panda PK;Van Tine BA
许多癌症使代谢酶乙酰氨基琥珀酸合成酶1（ASS 1）沉默，这是尿素循环中精氨酸生物合成的限速酶。因此，ASS 1阴性细胞对PEG化精氨酸脱亚胺酶（ADI-PEG 20）（目前正在临床试验中开发的一种药物）消耗细胞外精氨酸敏感。由于精氨酸耗竭抗性的主要机制是ASS 1的重新表达，我们寻求一种工具来理解单细胞水平上抗性表型的时间出现。开发了一种实时单细胞荧光生物传感器来监测精氨酸依赖性蛋白质翻译。这种多功能的基于蛋白质的传感器提供了有关细胞代谢适应的时间信息，因为它能够随着时间的推移量化和跟踪单个细胞。发现分析的每个ASS 1缺陷细胞通过传感器表达的减少对精氨酸剥夺作出反应，表明幼稚细胞群体中不存在抗性。然而，时间恢复和出现的电阻之间的细胞差异很大，这表明一个异质性的代谢反应。该传感器还能够确定其最佳翻译所需的最低精氨酸浓度。这里开发的依赖于抑制的传感器能够准确地跟踪用ADI-PEG 20处理的ASS 1缺陷细胞中的抗性的发展。其随时间推移追踪单细胞的能力允许确定在初始群体中不存在抗性，以及阐明抗性的时间和程度的异质性。该工具代表了精氨酸剥夺研究中的一个有用的进展，而其设计具有适应其他氨基酸的潜力。在线版本包含补充材料，可通过10.1186/s40170-021-00238-9获得。
Many cancers silence the metabolic enzyme argininosuccinate synthetase 1 (ASS1), the rate-limiting enzyme for arginine biosynthesis within the urea cycle. Consequently, ASS1-negative cells are susceptible to depletion of extracellular arginine by PEGylated arginine deiminase (ADI-PEG20), an agent currently being developed in clinical trials. As the primary mechanism of resistance to arginine depletion is re-expression of ASS1, we sought a tool to understand the temporal emergence of the resistance phenotype at the single-cell level. A real-time, single-cell florescence biosensor was developed to monitor arginine-dependent protein translation. The versatile, protein-based sensor provides temporal information about the metabolic adaptation of cells, as it is able to quantify and track individual cells over time. Every ASS1-deficient cell analyzed was found to respond to arginine deprivation by decreased expression of the sensor, indicating an absence of resistance in the naïve cell population. However, the temporal recovery and emergence of resistance varied widely amongst cells, suggesting a heterogeneous metabolic response. The sensor also enabled determination of a minimal arginine concentration required for its optimal translation. The translation-dependent sensor developed here is able to accurately track the development of resistance in ASS1-deficient cells treated with ADI-PEG20. Its ability to track single cells over time allowed the determination that resistance is not present in the naïve population, as well as elucidating the heterogeneity of the timing and extent of resistance. This tool represents a useful advance in the study of arginine deprivation, while its design has potential to be adapted to other amino acids. The online version contains supplementary material available at 10.1186/s40170-021-00238-9.
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影响因子：
8.8
作者：
Kremer JC;Prudner BC;Lange SES;Bean GR;Schultze MB;Brashears CB;Radyk MD;Redlich N;Tzeng SC;Kami K;Shelton L;Li A;Morgan Z;Bomalaski JS;Tsukamoto T;McConathy J;Michel LS;Held JM;Van Tine BA
通讯作者：
Van Tine BA
影响因子：
3.7
作者：
Kim JH;Lee SR;Li LH;Park HJ;Park JH;Lee KY;Kim MK;Shin BA;Choi SY
通讯作者：
Choi SY
影响因子：
6.2
作者：
Dillon, BJ;Prieto, VG;Clark, MA
通讯作者：
Clark, MA
影响因子：
14.9
作者：
Kaufman, Weimin L.;Kocman, Ibrahim;Agrawal, Vishal;Rahn, Hans-Peter;Besser, Daniel;Gossen, Manfred
通讯作者：
Gossen, Manfred
影响因子：
9
作者：
通讯作者：
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