Yeon Minjeong, Perego Michela, Elokely Khaled M, Abou-Gharbia Magid, Childers Wayne E, Milcarek Andrew T, Bertolini Irene, Tang Hsin-Yao, Languino Lucia R, Stein Gary S, Ghule Prachi N, Vietje Brad P, Taatjes Douglas T, Altieri Dario C
The Wistar Institute, Philadelphia, PA, United States.
University of Wyoming, Laramie, WY, United States.
Cancer Res. 2025 Sep 10. doi: 10.1158/0008-5472.CAN-25-0904.
Parkin is a mitochondria-associated E3 ubiquitin (Ub) ligase that mediates mitophagy and organelle quality control. More recently, Parkin has been implicated in stimulating antitumor immunity and reprogramming the tumor immune microenvironment. Here, we showed that Parkin ubiquitinates the alarmin molecule, high mobility group box-1 (HMGB1) on Lys146 (K146) using predominantly K48 linkages. By molecular modeling, the in-between-ring (IBR) domain of Parkin (Gln326-Leu358) made extensive contacts with the amino-terminus A box of HMGB1 (Met1-Ser42), forming a mitochondria-associated Parkin-HMGB1 complex that juxtaposes K146 to Ub active site residues Gly76 and Arg74. Instead of proteasomal degradation, Parkin ubiquitination of K146 enabled the loading of HMGB1, but not HMGB1 K146A mutant, onto autophagy- and mitochondria-derived large extracellular vesicles (LEV). In turn, released Parkin-HMGB1 LEV stimulated a potent interferon (IFN) and cytokine response in recipient cells, expanding CD8+ T cell subsets with effector (CD69+/KLRG1+), self-renewal (TCF-1+/PD-1+), and cytotoxic (KLRG1+/GrzB+) properties. Conditional expression of Parkin induced HMGB1 release, activated intratumoral CD8+ T cells, and suppressed syngeneic tumor growth in vivo in a response that was abolished by HMGB1 silencing. These data identify that Parkin-LEV regulated release of HMGB1 reprograms antitumor immunity via stimulation of IFN signaling and expansion of specialized CD8+ T cell subsets.
帕金蛋白是一种与线粒体相关的E3泛素连接酶，介导线粒体自噬和细胞器质量控制。最近，帕金蛋白被认为与刺激抗肿瘤免疫和重编程肿瘤免疫微环境有关。在此，我们发现帕金蛋白主要通过K48连接在赖氨酸146（K146）位点使警报素分子高迁移率族蛋白B1（HMGB1）发生泛素化。通过分子建模，帕金蛋白的环间（IBR）结构域（Gln326-Leu358）与HMGB1的氨基末端A盒（Met1-Ser42）广泛接触，形成一种与线粒体相关的帕金蛋白-HMGB1复合物，使K146与泛素活性位点残基Gly76和Arg74并列。帕金蛋白对K146的泛素化作用并非导致HMGB1被蛋白酶体降解，而是使HMGB1（而非HMGB1 K146A突变体）装载到自噬和线粒体来源的大型细胞外囊泡（LEV）上。反过来，释放的帕金蛋白-HMGB1 LEV在受体细胞中刺激了强烈的干扰素（IFN）和细胞因子反应，扩大了具有效应（CD69+/KLRG1+）、自我更新（TCF-1+/PD-1+）和细胞毒性（KLRG1+/GrzB+）特性的CD8+ T细胞亚群。帕金蛋白的条件性表达诱导HMGB1释放，激活肿瘤内CD8+ T细胞，并在体内抑制同基因肿瘤生长，而HMGB1沉默可消除这种反应。这些数据表明，帕金蛋白-LEV调节的HMGB1释放通过刺激IFN信号和扩大特定CD8+ T细胞亚群来重编程抗肿瘤免疫。
