ULK induces autophagy by targeting Beclin-1 in Vibrio splendidus challenged Apostichopus japonicus
ULK induces autophagy by targeting Beclin-1 in Vibrio splendidus challenged Apostichopus japonicus
复制标题
ULK 通过靶向 Beclin-1 诱导刺参攻击的灿烂弧菌自噬
DOI：
10.1016/j.aquaculture.2020.736022
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发表时间：
2021-02-15
期刊：
影响因子：
4.5
通讯作者：
Li, Chenghua
中科院分区：
文献类型：
--
作者：
Chen, Kaiyu;Shao, Yina;Li, Chenghua
ULK作为一种自噬诱导因子，在哺乳动物自噬的形成中起着至关重要的作用。然而，ULK对应物在棘皮动物自噬中的功能仍然不清楚。本研究利用RACE技术从仿刺参（Apostichopus）中克隆了一个新的ULK同源物，并对其生物学功能进行了研究。AjULK编码的蛋白含有DUF 3543（746-940个氨基酸）结构域和保守的丝氨酸/苏氨酸蛋白激酶（13-277个氨基酸）结构域。AjULK在所有检查的组织中广泛表达，在肌腱中发现最大的转录。在灿烂弧菌体内攻击后，AjULK和其他自噬相关基因，即AjAtg 13，AjBeclin-1和AjLC 3的mRNA水平显著上调。相反，在相同条件下，自噬底物Ajp 62的水平显著降低。亚细胞定位结果显示AjULK和AjAtg 13共定位于细胞质中，且感染后信号明显增强。结果表明，病原菌感染后，自噬被显著诱导。进一步的功能分析表明，与对照组相比，AjULK敲低后AjAtg 13和AjBeclin-1的蛋白水平同步下降。在相同条件下，Ajp 62蛋白水平显著上调。因此，在AjULK干扰后，自噬信号通过AjBeclin-1被显著抑制。AjULK干扰后细菌清除也明显受到抑制。因此，AjULK作为自噬启动子，通过靶向AjBeclin-1介导体腔细胞的自噬来限制细菌在海参中的入侵。
ULK serves as an autophagic inductor and plays crucial roles in the formation of autophagy in mammals. However, the function of the ULK counterpart in the autophagy of echinoderms remains ambiguous. In this study, a novel ULK homolog from the sea cucumber Apostichopus japonicus (AjULK) was cloned using RACE technology, and its biological function was investigated. AjULK encoded a predicted protein containing a DUF3543 (746-940 amino acids) domain and a conserved serine/threonine protein kinase (13-277 amino acids) domain. AjULK was widely expressed in all examined organizations, with the largest transcription found in the tentacle. After the Vibrio splendidus challenge in vivo, the mRNA levels of AjULK and other autophagy-related genes, namely, AjAtg13, AjBeclin-1, and AjLC3, were significantly upregulated. By contrast, the level of Ajp62, which was an autophagy substrate, was notably decreased under the same condition. Subcellular localization indicated that AjULK and AjAtg13 were colocalized in the cytoplasm, and the signal was significantly increased after V. splendidus infection. Result indicated that autophagy was markedly induced after pathogen infection. Further functional analysis showed that the protein levels of AjAtg13 and AjBeclin-1 were synchronously decreased after AjULK knockdown, compared with that of the control group. The protein level of Ajp62 was markedly upregulated under the same condition. Thus, the autophagy signal was significantly inhibited through AjBeclin-1 after AjULK interference. Bacterial clearance was also markedly inhibited after AjULK interference. Hence, AjULK served as an autophagic initiator by targeting AjBeclin-1 and mediating the autophagy of coelomocytes to restrict bacterial invasion in the sea cucumber.