Fu Chunmei, Wang Jie, Ma Tianle, Yin Congcong, Zhou Li, Clausen Björn E, Mi Qing-Sheng, Jiang Aimin
Center for Cutaneous Biology and Immunology, Department of Dermatology, Henry Ford Health, Detroit, MI 48202, USA.
Immunology Program, Henry Ford Cancer Institute, Henry Ford Health, Detroit, MI 48202, USA.
Vaccines (Basel). 2024 Sep 10;12(9):1037. doi: 10.3390/vaccines12091037.
GSK-3β plays a critical role in regulating the Wnt/β-catenin signaling pathway, and manipulating GSK-3β in dendritic cells (DCs) has been shown to improve the antitumor efficacy of DC vaccines. Since the inhibition of GSK-3β leads to the activation of β-catenin, we hypothesize that blocking GSK-3β in DCs negatively regulates DC-mediated CD8 T cell immunity and antitumor immunity. Using CD11c-GSK-3β conditional knockout mice in which GSK-3β is genetically deleted in CD11c-expressing DCs, we surprisingly found that the deletion of GSK-3β in DCs resulted in increased antitumor immunity, which contradicted our initial expectation of reduced antitumor immunity due to the presumed upregulation of β-catenin in DCs. Indeed, we found by both Western blot and flow cytometry that the deletion of GSK-3β in DCs did not lead to augmented expression of β-catenin protein, suggesting that GSK-3β exerts its function independent of β-catenin. Supporting this notion, our single-cell RNA sequencing (scRNA-seq) analysis revealed that GSK-3β-deficient DCs exhibited distinct gene expression patterns with minimally overlapping differentially expressed genes (DEGs) compared to DCs with activated β-catenin. This suggests that the deletion of GSK-3β in DCs is unlikely to lead to upregulation of β-catenin at the transcriptional level. Consistent with enhanced antitumor immunity, we also found that CD11c-GSK-3β mice exhibited significantly augmented cross-priming of antigen-specific CD8 T cells following DC-targeted vaccines. We further found that the deletion of GSK-3β in DCs completely abrogated memory CD8 T cell responses, suggesting that GSK-3β in DCs also plays a negative role in regulating the differentiation and/or maintenance of memory CD8 T cells. scRNA-seq analysis further revealed that although the deletion of GSK-3β in DCs positively regulated transcriptional programs for effector differentiation and function of primed antigen-specific CD8 T cells in CD11c-GSK-3β mice during the priming phase, it resulted in significantly reduced antigen-specific memory CD8 T cells, consistent with diminished memory responses. Taken together, our data demonstrate that GSK-3β in DCs has opposite functions in regulating cross-priming and memory CD8 T cell responses, and GSK-3β exerts its functions independent of its regulation of β-catenin. These novel insights suggest that targeting GSK-3β in cancer immunotherapies must consider its dual role in CD8 T cell responses.
糖原合成酶激酶-3β（GSK-3β）在调控Wnt/β-连环蛋白信号通路中起关键作用，并且已证明在树突状细胞（DC）中操控GSK-3β可提高DC疫苗的抗肿瘤疗效。由于抑制GSK-3β会导致β-连环蛋白激活，我们推测在DC中阻断GSK-3β会对DC介导的CD8 T细胞免疫和抗肿瘤免疫产生负向调节作用。利用CD11c-GSK-3β条件性敲除小鼠（其中在表达CD11c的DC中通过基因手段删除了GSK-3β），我们惊奇地发现DC中GSK-3β的缺失导致抗肿瘤免疫力增强，这与我们最初预期的由于DC中β-连环蛋白上调而导致抗肿瘤免疫力降低相矛盾。实际上，我们通过蛋白质免疫印迹法和流式细胞术均发现DC中GSK-3β的缺失并未导致β-连环蛋白蛋白表达增加，这表明GSK-3β发挥其功能独立于β-连环蛋白。支持这一观点的是，我们的单细胞RNA测序（scRNA-seq）分析显示，与β-连环蛋白激活的DC相比，缺乏GSK-3β的DC表现出独特的基因表达模式，差异表达基因（DEG）的重叠极少。这表明DC中GSK-3β的缺失不太可能在转录水平上导致β-连环蛋白上调。与增强的抗肿瘤免疫力一致，我们还发现CD11c-GSK-3β小鼠在接受DC靶向疫苗后，抗原特异性CD8 T细胞的交叉启动显著增强。我们进一步发现DC中GSK-3β的缺失完全消除了记忆性CD8 T细胞反应，这表明DC中的GSK-3β在调节记忆性CD8 T细胞的分化和/或维持方面也起负向作用。scRNA-seq分析进一步揭示，尽管在启动阶段CD11c-GSK-3β小鼠中DC中GSK-3β的缺失正向调节了已启动的抗原特异性CD8 T细胞效应分化和功能的转录程序，但它导致抗原特异性记忆性CD� T细胞显著减少，这与记忆反应减弱一致。综上所述，我们的数据表明DC中的GSK-3β在调节交叉启动和记忆性CD8 T细胞反应方面具有相反的功能，并且GSK-3β发挥其功能独立于其对β-连环蛋白的调节。这些新见解表明，在癌症免疫治疗中靶向GSK-3β必须考虑其在CD8 T细胞反应中的双重作用。
