Beclin 1 prevents ISG15-mediated cytokine storms to secure fetal hematopoiesis and survival

Proper control of inflammatory responses is essential for embryonic development, but the underlying mechanism is poorly understood. Here, we show that under physiological conditions, inactivation of ISG15, an inflammation amplifier, is associated with the interaction of Beclin 1 (Becn1), via its ECD domain, with STAT3 in the major fetal hematopoietic organ of mice. Conditional loss of Becn1 caused sequential dysfunction and exhaustion of fetal liver hematopoietic stem cells, leading to lethal inflammatory cell-biased hematopoiesis in the fetus. Molecularly, the absence of Becn1 resulted in the release of STAT3 from Becn1 tethering and subsequent phosphorylation and translocation to the nucleus, which in turn directly activated the transcription of ISG15 in fetal liver hematopoietic cells, coupled with increased ISGylation and production of inflammatory cytokines, whereas inactivating STAT3 reduced ISG15 transcription and inflammation but improved hematopoiesis potential, and further silencing ISG15 mitigated the above collapse in the Becn1 null hematopoietic lineage. The Becn1-STAT3-ISG15 axis remains functional in Atg5/7-disrupted fetal hematopoietic organs. These results suggest that Becn1, in an autophagy-independent manner, secures hematopoiesis and survival of the fetus by directly inhibiting STAT3-ISG15 activation to prevent cytokine storms. Our findings highlight a previously undocumented role of Becn1 in governing ISG15 to safeguard the fetus. Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for p-STAT3 (Tyr 705) in E14.5 Becn1-/- fetal livers.

炎症应答的适度调控对于胚胎发育不可或缺，但其潜在的分子机制至今尚未完全明晰。本研究显示，在生理状态下，作为炎症放大器的干扰素刺激基因15（ISG15）的失活，与小鼠主要胎儿造血器官中Beclin 1（Becn1）通过其ECD结构域（ECD domain）与信号转导与转录激活因子3（STAT3）的相互作用密切相关。条件性敲除Becn1会引发胎儿肝脏造血干细胞（hematopoietic stem cells）的进行性功能障碍与耗竭，最终导致胎儿出现以炎症细胞为优势的致死性造血异常。从分子层面来看，Becn1的缺失会使STAT3脱离其束缚，随后发生磷酸化并转位入核，进而直接激活胎儿肝脏造血细胞中ISG15的转录，同时伴随ISG化修饰（ISGylation）水平升高与炎症细胞因子生成增多；而抑制STAT3活性则可降低ISG15的转录与炎症反应，但能提升造血潜能，进一步沉默ISG15则可缓解Becn1缺失型造血谱系中的上述异常崩溃。在自噬相关基因Atg5/7敲除的胎儿造血器官中，Becn1-STAT3-ISG15调控轴仍可正常发挥功能。上述结果表明，Becn1可不依赖自噬的方式，通过直接抑制STAT3-ISG15通路的激活以阻断细胞因子风暴，从而维持胎儿的造血功能与存活。本研究揭示了Becn1此前未被报道的调控ISG15的功能，从而为胎儿存活提供保障。本数据集包含针对E14.5天Becn1基因敲除胎鼠肝脏中磷酸化STAT3（Tyr705，p-STAT3）的染色质免疫沉淀测序（Chromatin immunoprecipitation DNA-sequencing，ChIP-seq）数据。