DataSheet1_Proliferation and migration of ML1 follicular thyroid cancer cells are inhibited by IU1 targeting USP14: role of proteasome and autophagy flux.PDF

USP14 is a deubiquitinating enzyme involved in protein degradation by interacting with the proteasome and removal of poly-ubiquitin chains on target proteins. USP14 can influence cellular processes such as cell survival, DNA repair, ER stress, endocytosis, and the inflammatory response. USP14 further plays a role in tumor growth, and the inhibition of USP14 by compounds such as IU1 may affect cancer cell migration and invasion. Here we have studied the mechanisms for the action of IU1 in ML1 follicular thyroid cancer cells, comparing them with control, primary thyroid cells. Treatment with IU1 reduced proliferation of ML1 cells in a concentration-dependent manner, and more prominently than in control cells. IU1 decreased basal migration of ML1 cells, and after stimulation of cells with the bioactive compound, sphingosine-1-phosphate. The sphingosine-1-phosphate receptor 3 was increased in ML1 cells as compared with control thyroid cells, but this was not influenced by IU1. Further studies on the mechanism, revealed that IU1 enhanced the proteasome activity as well as LC3B-dependent autophagy flux in ML1 cells with an opposite effect on control thyroid cells. This indicates that IU1 elicits a cell-type dependent autophagy response, increasing it in ML1 cancer cells. The IU1-mediated stimulation of autophagy and proteasomes can likely contribute to the reduced cell proliferation and migration observed in ML1 cells. The precise set of proteins affected by IU1 in ML1 thyroid and other cancer cells warrant further investigations.

USP14是一种去泛素化酶（deubiquitinating enzyme），可通过与蛋白酶体（proteasome）相互作用、去除靶蛋白的多聚泛素链（poly-ubiquitin chains）参与蛋白质降解过程。USP14能够调控细胞存活（cell survival）、DNA修复（DNA repair）、内质网应激（ER stress）、胞吞作用（endocytosis）及炎症反应（inflammatory response）等多种细胞生命活动。此外，USP14还参与肿瘤生长（tumor growth）过程；而IU1等化合物对USP14的抑制作用，可影响癌细胞的迁移与侵袭能力。本研究以ML1滤泡状甲状腺癌细胞（ML1 follicular thyroid cancer cells）为对象，探究IU1的作用机制，并与对照组原代甲状腺细胞（primary thyroid cells）开展对照分析。经IU1处理后，ML1细胞的增殖能力呈浓度依赖性降低，且该抑制效果相较于对照组原代甲状腺细胞更为显著。IU1可降低ML1细胞的基础迁移能力，同时也能削弱经生物活性化合物鞘氨醇-1-磷酸（sphingosine-1-phosphate）刺激后的细胞迁移能力。相较于对照组原代甲状腺细胞，ML1细胞中鞘氨醇-1-磷酸受体3（sphingosine-1-phosphate receptor 3）的表达水平更高，但该表达差异并未受到IU1的调控。进一步的机制研究表明，IU1可增强ML1细胞的蛋白酶体活性与LC3B依赖性自噬流（LC3B-dependent autophagy flux），而该作用与对照组原代甲状腺细胞中的效应恰好相反。这一结果提示，IU1可引发细胞类型依赖性的自噬应答：在ML1癌细胞中，IU1能够促进自噬过程。IU1介导的自噬激活与蛋白酶体活性增强，或可解释ML1细胞中观察到的细胞增殖与迁移能力下降现象。目前，IU1在ML1甲状腺癌细胞及其他癌细胞中所影响的具体蛋白谱系，仍有待进一步深入研究。