生长素诱导的降解结构域 2 (AID2) 系统可在秀丽隐杆线虫的胚胎发生和发育过程中实现对蛋白质的可控敲低。
The auxin-inducible degron 2 (AID2) system enables controlled protein knockdown during embryogenesis and development in Caenorhabditis elegans.
机构信息
Multicellular Organization Laboratory, Department of Gene Function and Phenomics, National Institute of Genetics, Research Organization of Information and Systems (ROIS), Mishima, Shizuoka 411-8540, Japan.
Department of Genetics, School of Life Science, SOKENDAI (The Graduate University for Advanced Studies), Mishima, Shizuoka 411-8540, Japan.
出版信息
Genetics. 2022 Feb 4;220(2). doi: 10.1093/genetics/iyab218.
Targeted protein degradation using the auxin-inducible degron (AID) system is garnering attention in the research field of Caenorhabditis elegans, because of the rapid and efficient target depletion it affords, which can be controlled by treating the animals with the phytohormone auxin. However, the current AID system has drawbacks, i.e., leaky degradation in the absence of auxin and the requirement for high auxin doses. Furthermore, it is challenging to deplete degron-fused proteins in embryos because of their eggshell, which blocks auxin permeability. Here, we apply an improved AID2 system utilizing AtTIR1(F79G) and 5-phenyl-indole-3-acetic acid (5-Ph-IAA) to C. elegans and demonstrated that it confers better degradation control vs the previous system by suppressing leaky degradation and inducing sharp degradation using 1,300-fold lower 5-Ph-IAA doses. We successfully degraded the endogenous histone H2A.Z protein fused to an mAID degron and disclosed its requirement in larval growth and reproduction, regardless of the presence of maternally inherited H2A.Z molecules. Moreover, we developed an eggshell-permeable 5-Ph-IAA analog, 5-Ph-IAA-AM, that affords an enhanced degradation in laid embryos. Our improved system will contribute to the disclosure of the roles of proteins in C. elegans, in particular those that are involved in embryogenesis and development, through temporally controlled protein degradation.
利用生长素诱导的降解(AID)系统进行靶向蛋白降解在秀丽隐杆线虫的研究领域引起了关注,因为它提供了快速和有效的靶标消耗,可以通过用植物激素生长素处理动物来控制。然而,目前的 AID 系统存在一些缺点,即在没有生长素的情况下存在漏降解,并且需要高剂量的生长素。此外,由于卵壳阻止了生长素的通透性,因此在胚胎中耗尽融合了降解结构域的蛋白质具有挑战性。在这里,我们应用了一种利用 AtTIR1(F79G)和 5-苯基吲哚-3-乙酸(5-Ph-IAA)的改良 AID2 系统来处理秀丽隐杆线虫,并证明与之前的系统相比,它通过抑制漏降解并使用 1300 倍低剂量的 5-Ph-IAA 诱导急剧降解,提供了更好的降解控制。我们成功地降解了融合到 mAID 降解结构域的内源组蛋白 H2A.Z 蛋白,并揭示了其在幼虫生长和繁殖中的需求,而与母体遗传的 H2A.Z 分子是否存在无关。此外,我们开发了一种卵壳通透性的 5-Ph-IAA 类似物 5-Ph-IAA-AM,它可以在产卵的胚胎中提供增强的降解。我们的改良系统将通过时间控制的蛋白质降解,有助于揭示秀丽隐杆线虫中蛋白质的作用,特别是那些参与胚胎发生和发育的蛋白质的作用。