Ulas Evgeniia, Brodsky Ilya, Burakov Anton
A.N. Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow 119992, Russia.
Institute of Protein Research of Russian Academy of Sciences, Pushchino 142290, Russia.
Biomedicines. 2024 Dec 18;12(12):2872. doi: 10.3390/biomedicines12122872.
The changes in the level of small GTPase ARL4C are associated with the initiation and progression of many different cancers. The content of ARL4C varies greatly between different tissues, and the induction of ARL4C expression leads to changes in cell morphology and proliferation. Although ARL4C can bind alpha-tubulin and affect intracellular transport, the role of ARL4C in the functioning of the tubulin cytoskeleton remained unclear. The aim of the present work is to study this role; Methods: The cells of the following lines were used for the experiments: HeLa (human cervical carcinoma), MCF7 (human breast cancer), U2OS (human osteosarcoma), Vero, BS-C-1, and COS7 (African green monkey kidney). The receptor activation by agonists followed by the preparation of cell lysates, electrophoresis, and immunoblotting, as well as cell fixation and immunofluorescent staining, were used to assess endogenous ARL4C/ABCA1 levels and the microtubule network morphology. The microtubule regrowth technique was performed to estimate the rate of microtubule nucleation, and the overexpression of different ARL4C constructs was used to affect ARL4C activity in the cells; Results: We showed that the changes in the endogenous ARL4C level or the ARL4C activity alter the microtubule nucleation process in the cells; Conclusions: small GTPase ARL4C may serve as one of the regulators of the microtubule nucleation process both in normal and cancer cells.
小GTP酶ARL4C水平的变化与许多不同癌症的发生和发展相关。ARL4C的含量在不同组织之间差异很大,ARL4C表达的诱导会导致细胞形态和增殖的变化。尽管ARL4C可以结合α-微管蛋白并影响细胞内运输,但ARL4C在微管蛋白细胞骨架功能中的作用仍不清楚。本研究的目的是探讨这一作用;方法:使用以下细胞系进行实验:HeLa(人宫颈癌)、MCF7(人乳腺癌)、U2OS(人骨肉瘤)、Vero、BS-C-1和COS7(非洲绿猴肾)。通过激动剂激活受体,随后制备细胞裂解物、进行电泳和免疫印迹,以及细胞固定和免疫荧光染色,来评估内源性ARL4C/ABCA1水平和微管网络形态。采用微管再生技术来估计微管成核速率,并使用不同的ARL4C构建体的过表达来影响细胞中的ARL4C活性;结果:我们发现内源性ARL4C水平或ARL4C活性的变化会改变细胞中的微管成核过程;结论:小GTP酶ARL4C可能是正常细胞和癌细胞中微管成核过程的调节因子之一。