García-Hernández Brenda, Morán Julio
División de Neurociencias, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, Mexico City, Mexico.
Front Mol Neurosci. 2023 Jul 20;16:1210962. doi: 10.3389/fnmol.2023.1210962. eCollection 2023.
TXNIP is a protein sensitive to oxidant conditions whose expression is related to the progression of death in cancer, diabetes, ischemia, and neurodegenerative diseases, among others. Because of this, many studies propose TXNIP as a therapeutic target in several diseases. Exposure of cerebellar granule neurons to staurosporine or low potassium leads to apoptotic death. Both conditions generate an early production of reactive oxygen species (ROS) that induces the activation of the ASK1 pathway and the apoptotic machinery. In these models, it has been shown an increase in TXNIP protein mediated by ROS. Here, we evaluated the molecular mechanisms involved in the regulation of the expression during neuronal death, as well as the role of the protein in the progression of cell death induced by these two apoptotic conditions. In cultured cerebellar granule neurons, we observed that low potassium and staurosporine induced an early increase in ROS that correlated with an increase in mRNA. When we evaluated the promoter of the gene, we found that the JASPAR-reported FOXO1/3 transcription factor motifs are close to the transcription start site (TSS). We then verified through the Chromatin immunoprecipitation technique (ChIP) that FOXO3 interacts with the promoter after 1 h of low potassium treatment. We also detected FOXO3 nuclear translocation by low potassium and staurosporine treatments. Finally, by using shRNA in the neuroblastoma MSN cell line, we found that downregulation decreased neuronal death induced by staurosporine stimulus. Together, these results suggest that ROS promotes the expression of through the activation of the FOXO3 transcription factor mediated by Akt inhibition. We also demonstrated that TXNIP is necessary for neuronal death progression.
硫氧还蛋白相互作用蛋白（TXNIP）是一种对氧化条件敏感的蛋白质，其表达与癌症、糖尿病、缺血和神经退行性疾病等多种疾病的死亡进程相关。正因如此，许多研究提出将TXNIP作为多种疾病的治疗靶点。小脑颗粒神经元暴露于星形孢菌素或低钾环境会导致凋亡性死亡。这两种情况都会早期产生活性氧（ROS），从而诱导ASK1信号通路和凋亡机制的激活。在这些模型中，已显示ROS介导TXNIP蛋白增加。在此，我们评估了神经元死亡过程中参与其表达调控的分子机制，以及该蛋白在这两种凋亡条件诱导的细胞死亡进程中的作用。在培养的小脑颗粒神经元中，我们观察到低钾和星形孢菌素会诱导ROS早期增加，这与TXNIP mRNA增加相关。当我们评估该基因的启动子时，发现JASPAR报告的FOXO1/3转录因子基序靠近转录起始位点（TSS）。然后我们通过染色质免疫沉淀技术（ChIP）验证了低钾处理1小时后FOXO3与TXNIP启动子相互作用。我们还通过低钾和星形孢菌素处理检测到FOXO3的核转位。最后，通过在神经母细胞瘤MSN细胞系中使用短发夹RNA（shRNA），我们发现TXNIP下调可减少星形孢菌素刺激诱导的神经元死亡。综上所述，这些结果表明ROS通过抑制Akt介导的FOXO3转录因子激活促进TXNIP的表达。我们还证明了TXNIP是神经元死亡进程所必需的。
