Embryonic Cells Redistribute SUMO1 upon Forced SUMO1 Overexpression
Embryonic Cells Redistribute SUMO1 upon Forced SUMO1 Overexpression
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DOI：
10.1128/mbio.01856-19
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发表时间：
2019-11-01
期刊：
影响因子：
6.4
通讯作者：
Goff, Stephen P.
中科院分区：
文献类型：
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作者：
Lee, Andreia;Zhu, Yiping;Goff, Stephen P.
小泛素样修饰物（SUMO）与底物蛋白的缀合是一种翻译后蛋白质修饰，影响多种生理过程。小鼠SUMO结合的全面抑制导致胚胎死亡，反映了SUMO途径对胚胎发育的重要性。在这里，我们证明了SUMO 1过表达在小鼠胚胎癌和胚胎干（ES）细胞中耐受性不好，并且在用递送SUMO 1表达构建体的载体转导后仅回收了少数克隆。分化的NIH/3 T3细胞过度表达SUMO 1而没有有害作用，并保持高水平的结合和游离形式的SUMO 1。在强制过表达后存活的少数胚胎细胞以少数高分子量缀合物的形式保留了所有的SUMO 1，并保持了不可检测的游离SUMO 1水平。胚胎细胞中游离SUMO的缺乏在SUMO 1过表达时特异性地观察到，但SUMO 2没有。此外，通过SUMO 1的C-末端突变或通过SUMO 1底物“海绵”的过表达或通过去SUMO化酶SUMO特异性肽酶1（SENP 1）的过表达来阻断SUMO 1与内源性底物的缀合显著地恢复了游离SUMO 1的过表达。这些数据表明，SUMO 1蛋白的过度表达导致关键SUMO 1结合底物的过量积累在胚胎细胞中是不耐受的。存活的胚胎细胞表现出SUMO 1结合到允许的底物上，但完全不存在游离的SUMO 1。重要胚胎干细胞（ES）表现出不寻常的转录、蛋白质组和信号反应谱，反映了它们对快速分化和复制的不寻常需求。本文报道的工作表明，小鼠胚胎细胞系不能耐受SUMO 1的过表达，SUMO 1是一种小的泛素样修饰蛋白，共价连接到许多底物上，以改变其细胞内定位和功能。SUMO 1的过度表达对ES细胞是有毒的，存活的细胞群体通过显著降低游离SUMO 1的水平来适应。在分化的细胞中，或在SUMO 2或非缀合的SUMO 1突变体中，或在接受修饰的SUMO 1“海绵”底物存在下，均未观察到这种反应。这些发现表明，胚胎细胞不能耐受特定底物的过量SUMO 1修饰，并强调了这些细胞控制可用于缀合的SUMO 1水平的独特需求。
Conjugation of small ubiquitin-like modifiers (SUMOs) to substrate proteins is a posttranslational protein modification that affects a diverse range of physiological processes. Global inhibition of SUMO conjugation in mice results in embryonic lethality, reflecting the importance of the SUMO pathways for embryonic development. Here, we demonstrated that SUMO1 overexpression was not well tolerated in murine embryonic carcinoma and embryonic stem (ES) cells and that only a few clones were recovered after transduction with vectors delivering SUMO1 expression constructs. Differentiated NIH/3T3 cells overexpress SUMO1 without deleterious effects and maintain high levels of both conjugated and free forms of SUMO1. The few embryonic cells surviving after forced overexpression retained all their SUMO1 in the form of a few high-molecular-weight conjugates and maintained undetectable levels of free SUMO1. The absence of free SUMO in embryonic cells was seen specifically upon overexpression of SUMO1, but not SUMO2. Moreover, blocking SUMO1 conjugation to endogenous substrates by C-terminal mutations of SUMO1 or by overexpression of a SUMO1 substrate "sponge" or by overexpression of the deSUMOylating enzyme SUMO-specific peptidase 1 (SENP1) dramatically restored free SUMO1 overexpression. The data suggest that overexpression of SUMO1 protein leading to an excess accumulation of critical SUMO1-conjugated substrates is not tolerated in embryonic cells. Surviving embryonic cells exhibit SUMO1 conjugation to allowed substrates but a complete absence of free SUMO1.IMPORTANCE Embryonic stem ( ES) cells exhibit unusual transcriptional, proteomic, and signal response profiles, reflecting their unusual needs for rapid differentiation and replication. The work reported here demonstrated that mouse embryonic cell lines did not tolerate the overexpression of SUMO1, the small ubiquitin-like modifier protein that is covalently attached to many substrates to alter their intracellular localization and functionality. Forced SUMO1 overexpression is toxic to ES cells, and surviving cell populations adapt by dramatically reducing the levels of free SUMO1. Such a response is not seen in differentiated cells or with SUMO2 or with nonconjugatable SUMO1 mutants or in the presence of a SUMO1 "sponge" substrate that accepts the modification. The findings suggest that excess SUMO1 modification of specific substrates is not tolerated by embryonic cells and highlight a distinctive need for these cells to control the levels of SUMO1 available for conjugation.