Necroptosis is emerging as a new target for cancer immunotherapy as it is now recognized as a form of cell death that increases tumor immunogenicity, which would be especially helpful in treating immune-desert tumors. De novo synthesis of inflammatory proteins during necroptosis appears especially important in facilitating increased anti-tumor immune responses. While late-stage transcription mediated by NF-κB during cell death is believed to play a role in this process, it is otherwise unclear what cell signaling events initiate this transactivation of inflammatory genes. We employed tandem-affinity purification linked to mass spectrometry (TAP-MS), in combination with the analysis of RNA-sequencing (RNA-Seq) datasets to identify the Tripartite Motif Protein 28 (TRIM28) as a candidate co-repressor. Comprehensive biochemical and molecular biology techniques were used to characterize the role of TRIM28 in RIPK3 activation-induced transcriptional and immunomodulatory events. The cell composition estimation module was used to evaluate the correlation between RIPK3/TRIM28 levels and CD8+ T cells or dendritic cells (DC) in all TCGA tumors. We identified TRIM28 as a co-repressor that regulates transcriptional activity during necroptosis. Activated RIPK3 phosphorylates TRIM28 on serine 473, inhibiting its chromatin binding activity, thereby contributing to the transactivation of NF-κB and other transcription factors, such as SOX9. This leads to elevated cytokine expression, which then potentiates immunoregulatory processes, such as DC maturation. The expression of RIPK3 has a significant positive association with the tumor-infiltrating immune cells populations in various tumor type, thereby activating anti-cancer responses. Our data suggest that RIPK3 activation-dependent derepression of TRIM28 in cancer cells leads to increased immunostimulatory cytokine production in the tumor microenvironment, which then contributes to robust cytotoxic anti-tumor immunity.

中文翻译：

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RIPK3激活诱导癌细胞中TRIM28去抑制并增强抗肿瘤微环境

坏死性凋亡正在成为癌症免疫治疗的新靶点，因为它现在被认为是一种增加肿瘤免疫原性的细胞死亡形式，这将特别有助于治疗免疫沙漠肿瘤。坏死性凋亡过程中炎症蛋白的从头合成在促进抗肿瘤免疫反应增强方面显得尤为重要。虽然据信在细胞死亡期间由 NF-κB 介导的晚期转录在此过程中发挥作用，但尚不清楚是哪些细胞信号传导事件启动了炎症基因的这种反式激活。我们采用与质谱 (TAP-MS) 相关的串联亲和纯化，结合 RNA 测序 (RNA-Seq) 数据集的分析来确定三方基序蛋白 28 (TRIM28) 作为候选共阻遏物。综合生化和分子生物学技术用于表征 TRIM28 在 RIPK3 激活诱导的转录和免疫调节事件中的作用。细胞组成估计模块用于评估所有 TCGA 肿瘤中 RIPK3/TRIM28 水平与 CD8+ T 细胞或树突状细胞 (DC) 之间的相关性。我们将 TRIM28 鉴定为在坏死性凋亡过程中调节转录活性的共抑制因子。活化的 RIPK3 磷酸化丝氨酸 473 上的 TRIM28，抑制其染色质结合活性，从而促进 NF-κB 和其他转录因子（如 SOX9）的反式激活。这导致细胞因子表达升高，然后增强免疫调节过程，例如 DC 成熟。RIPK3的表达与各种肿瘤类型的肿瘤浸润性免疫细胞群呈显着正相关，从而激活抗癌反应。我们的数据表明，癌细胞中TRIM28的RIPK3激活依赖性去抑制导致肿瘤微环境中免疫刺激性细胞因子的产生增加，从而有助于产生强大的细胞毒性抗肿瘤免疫。