Borkúti Péter, Kristó Ildikó, Szabó Anikó, Bajusz Csaba, Kovács Zoltán, Réthi-Nagy Zsuzsánna, Lipinszki Zoltán, Lukácsovich Tamás, Bogdan Sven, Vilmos Péter
Eötvös Loránd Research Network (ELKH), Biological Research Centre, Szeged, Hungary.
Doctoral School of Multidisciplinary Medical Science, University of Szeged, Szeged, Hungary.
Front Mol Biosci. 2022 Aug 19;9:963635. doi: 10.3389/fmolb.2022.963635. eCollection 2022.
Actin, as an ancient and fundamental protein, participates in various cytoplasmic as well as nuclear functions in eukaryotic cells. Based on its manifold tasks in the nucleus, it is a reasonable assumption that the nuclear presence of actin is essential for the cell, and consequently, its nuclear localization is ensured by a robust system. However, today only a single nuclear import and a single nuclear export pathway is known which maintain the dynamic balance between cytoplasmic and nuclear actin pools. In our work, we tested the robustness of the nuclear import of actin, and investigated whether the perturbations of nuclear localization affect the viability of the whole organism. For this aim, we generated a genetic system in in which we rescued the lethal phenotype of the null mutation of the gene with transgenes that express different derivatives of actin, including a Nuclear Export Signal (NES)-tagged isoform which ensures forced nuclear export of the protein. We also disrupted the SUMOylation site of actin, suggested earlier to be responsible for nuclear retention, and eliminated the activity of the single nuclear import factor dedicated to actin. We found that, individually, none of the above mentioned manipulations led to a notable reduction in nuclear actin levels and thus, fully rescued lethality. However, the NES tagging of actin, together with the knock out of its importin, significantly reduced the amount of nuclear actin and induced lethality, confirming that the presence of actin in the nucleus is essential, and thereby, over-secured. Supporting this, we identified novel nuclear importins specific to actin, which sheds light on the mechanism behind the robustness of nuclear localization of actin, and supports the idea of essentiality of its nuclear functions.
肌动蛋白作为一种古老而基础的蛋白质，参与真核细胞的各种细胞质及细胞核功能。基于其在细胞核中的多种任务，可以合理推测肌动蛋白在细胞核中的存在对细胞至关重要，因此，其核定位由一个强大的系统来确保。然而，如今已知的维持细胞质和细胞核肌动蛋白池之间动态平衡的核输入和核输出途径各只有一条。在我们的研究中，我们测试了肌动蛋白核输入的稳健性，并研究了核定位的扰动是否会影响整个生物体的生存能力。为此，我们构建了一个遗传系统，在该系统中，我们用表达肌动蛋白不同衍生物的转基因来挽救基因敲除突变的致死表型，这些衍生物包括一个带有核输出信号（NES）标签的异构体，可确保该蛋白的强制核输出。我们还破坏了先前认为负责核滞留的肌动蛋白的SUMO化位点，并消除了专门负责肌动蛋白的单核输入因子的活性。我们发现，上述操作单独进行时，均未导致核肌动蛋白水平显著降低，因此完全挽救了致死性。然而，肌动蛋白的NES标签与敲除其输入蛋白一起，显著降低了核肌动蛋白的量并诱导了致死性，这证实了肌动蛋白在细胞核中的存在至关重要，从而得到了过度保障。支持这一点的是，我们鉴定出了肌动蛋白特有的新型核输入蛋白，这揭示了肌动蛋白核定位稳健性背后的机制，并支持了其核功能至关重要的观点。
