STK3 kinase activation inhibits tumor proliferation through FOXO1-TP53INP1/P21 pathway in esophageal squamous cell carcinoma.

IF 4.9 2区 医学 Q2 CELL BIOLOGY Cellular Oncology Pub Date : 2024-08-01 Epub Date: 2024-03-04 DOI:10.1007/s13402-024-00928-8
Ziying Zhao, Yuan Chu, Anqi Feng, Shihan Zhang, Hao Wu, Zhaoxing Li, Mingchuang Sun, Li Zhang, Tao Chen, Meidong Xu
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Abstract

Purpose: Esophageal squamous cell carcinoma (ESCC) is an aggressive disease with a poor prognosis, caused by the inactivation of critical cell growth regulators that lead to uncontrolled proliferation and increased malignancy. Although Serine/Threonine Kinase 3 (STK3), also known as Mammalian STE20-like protein kinase 2 (MST2), is a highly conserved kinase of the Hippo pathway, plays a critical role in immunomodulation, organ development, cellular differentiation, and cancer suppression, its phenotype and function in ESCC require further investigation. In this study, we report for the first time on the role of STK3 kinase and its activation condition in ESCC, as well as the mechanism and mediators of kinase activation.

Methods: In this study, we investigated the expression and clinical significance of STK3 in ESCC. We first used bioinformatics databases and immunohistochemistry to analyze STK3 expression in the ESCC patient cohort and conducted survival analysis. In vivo, we conducted a tumorigenicity assay using nude mouse models to demonstrate the phenotypes of STK3 kinase. In vitro, we conducted Western blot analysis, qPCR analysis, CO-IP, and immunofluorescence (IF) staining analysis to detect molecule expression, interaction, and distribution. We measured proliferation, migration, and apoptosis abilities in ESCC cells in the experimental groups using CCK-8 and transwell assays, flow cytometry, and EdU staining. We used RNA-seq to identify genes that were differentially expressed in ESCC cells with silenced STK3 or FOXO1. We demonstrated the regulatory relationship of the TP53INP1/P21 gene medicated by the STK3-FOXO1 axis using Western blotting and ChIP in vitro.

Results: We demonstrate high STK3 expression in ESCC tissue and cell lines compared to esophageal epithelium. Cellular ROS induces STK3 autophosphorylation in ESCC cells, resulting in upregulated p-STK3/4. STK3 activation inhibits ESCC cell proliferation and migration by triggering apoptosis and suppressing the cell cycle. STK3 kinase activation phosphorylates FOXO1Ser212, promoting nuclear translocation, enhancing transcriptional activity, and upregulating TP53INP1 and P21. We also investigated TP53INP1 and P21's phenotypic effects in ESCC, finding that their knockdown significantly increases tumor proliferation, highlighting their crucial role in ESCC tumorigenesis.

Conclusion: STK3 kinase has a high expression level in ESCC and can be activated by cellular ROS, inhibiting cell proliferation and migration. Additionally, STK3 activation-mediated FOXO1 regulates ESCC cell apoptosis and cell cycle arrest by targeting TP53INP1/P21. Our research underscores the anti-tumor function of STK3 in ESCC and elucidates the mechanism underlying its anti-tumor effect on ESCC.

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STK3 激酶激活通过 FOXO1-TP53INP1/P21 通路抑制食管鳞状细胞癌的肿瘤增殖
目的:食管鳞状细胞癌(ESCC)是一种预后不良的侵袭性疾病,其病因是关键细胞生长调节因子失活,导致增殖失控和恶性程度增加。虽然丝氨酸/苏氨酸激酶 3(STK3),又称哺乳动物 STE20 样蛋白激酶 2(MST2),是一种高度保守的 Hippo 通路激酶,在免疫调节、器官发育、细胞分化和癌症抑制中发挥着关键作用,但它在 ESCC 中的表型和功能还需要进一步研究。本研究首次报道了STK3激酶在ESCC中的作用及其激活条件,以及激酶激活的机制和介质:本研究探讨了 STK3 在 ESCC 中的表达及其临床意义。我们首先利用生物信息学数据库和免疫组化技术分析了 STK3 在 ESCC 患者队列中的表达情况,并进行了生存分析。在体内,我们利用裸鼠模型进行了致瘤性试验,以证明 STK3 激酶的表型。在体外,我们进行了 Western 印迹分析、qPCR 分析、CO-IP 和免疫荧光(IF)染色分析,以检测分子的表达、相互作用和分布。我们使用 CCK-8 和 transwell 试验、流式细胞仪和 EdU 染色法检测了实验组 ESCC 细胞的增殖、迁移和凋亡能力。我们使用 RNA-seq 鉴定了 STK3 或 FOXO1 沉默的 ESCC 细胞中差异表达的基因。我们利用 Western 印迹技术和体外 ChIP 技术证明了 STK3-FOXO1 轴对 TP53INP1/P21 基因的调控关系:结果:与食管上皮细胞相比,我们发现 STK3 在 ESCC 组织和细胞系中高表达。细胞ROS诱导ESCC细胞中STK3自身磷酸化,导致p-STK3/4上调。STK3 激活可通过触发细胞凋亡和抑制细胞周期来抑制 ESCC 细胞的增殖和迁移。STK3 激酶活化可使 FOXO1Ser212 磷酸化,促进核转位,增强转录活性,上调 TP53INP1 和 P21。我们还研究了TP53INP1和P21在ESCC中的表型效应,发现敲除它们会显著增加肿瘤的增殖,突出了它们在ESCC肿瘤发生中的关键作用:结论:STK3激酶在ESCC中表达水平较高,可被细胞ROS激活,抑制细胞增殖和迁移。此外,STK3 激活介导的 FOXO1 通过靶向 TP53INP1/P21 调节 ESCC 细胞凋亡和细胞周期停滞。我们的研究强调了 STK3 在 ESCC 中的抗肿瘤功能,并阐明了其对 ESCC 抗肿瘤作用的机制。
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来源期刊
Cellular Oncology
Cellular Oncology ONCOLOGY-CELL BIOLOGY
CiteScore
10.30
自引率
1.50%
发文量
86
审稿时长
12 months
期刊介绍: The Official Journal of the International Society for Cellular Oncology Focuses on translational research Addresses the conversion of cell biology to clinical applications Cellular Oncology publishes scientific contributions from various biomedical and clinical disciplines involved in basic and translational cancer research on the cell and tissue level, technical and bioinformatics developments in this area, and clinical applications. This includes a variety of fields like genome technology, micro-arrays and other high-throughput techniques, genomic instability, SNP, DNA methylation, signaling pathways, DNA organization, (sub)microscopic imaging, proteomics, bioinformatics, functional effects of genomics, drug design and development, molecular diagnostics and targeted cancer therapies, genotype-phenotype interactions. A major goal is to translate the latest developments in these fields from the research laboratory into routine patient management. To this end Cellular Oncology forms a platform of scientific information exchange between molecular biologists and geneticists, technical developers, pathologists, (medical) oncologists and other clinicians involved in the management of cancer patients. In vitro studies are preferentially supported by validations in tumor tissue with clinicopathological associations.
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