<br>SPI1 通过 AKT1/mTOR 通路调节 HK2 参与恶性黑色素瘤发病机制,Journal of Cellular and Molecular Medicine

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SPI1 通过 AKT1/mTOR 通路调节 HK2 参与恶性黑色素瘤发病机制
Journal of Cellular and Molecular Medicine ( IF 4.3 ) Pub Date : 2023-08-04 , DOI: 10.1111/jcmm.17844
Chunlei Liu ₁ , Xiujuan Qiu ₂ , Jun Gao ₃ , Zhifan Gong ₁ , Xiaogang Zhou ₁ , Haichao Luo ₂ , Xuerui Geng ₁

Affiliation

Spi-1原癌基因（SPI1）在致癌过程中发挥着至关重要的作用。我们的工作旨在研究 SPI1 在黑色素瘤中的潜在调控机制。通过 qRT-PCR 和蛋白质印迹法测量 mRNA 和蛋白质水平。通过CCK-8测定评估细胞活力。使用双荧光素酶报告基因检测确定 SPI1 和己糖激酶 2 (HK2) 之间的靶标关系。进行 ChIP 是为了确认 SPI1 和 HK2 启动子之间的靶向关系。免疫组织化学分析检测黑色素瘤组织中SPI1和HK2的阳性细胞数。使用伤口愈合测定来确定细胞迁移能力。测量葡萄糖消耗、丙酮酸脱氢酶活性、乳酸产生和 ATP 水平以评估糖酵解。黑色素瘤细胞和组织中的 SPI1 转录分别显着高于邻近正常组织和表皮黑色素细胞 HEMa-LP。 SPI1 的敲低抑制了黑色素瘤细胞的细胞活力、转移和糖酵解。 SPI1直接靶向HK2，敲除SPI1可抑制HK2表达。 HK2的过表达削弱了SPI1敲低对黑色素瘤细胞的活力、转移和糖酵解的抑制作用。丝氨酸-苏氨酸激酶 1 (AKT1)/哺乳动物雷帕霉素靶蛋白 (mTOR) 轴参与黑色素瘤进展。 SPI1 敲低通过调节 AKT1/mTOR 通路抑制黑色素瘤细胞增殖、转移和糖酵解。

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SPI1 involvement in malignant melanoma pathogenesis by regulation of HK2 through the AKT1/mTOR pathway

Spi-1 proto-oncogene (SPI1) plays a vital role in carcinogenesis. Our work aimed to investigate the potential regulatory mechanism of SPI1 in melanoma. The mRNA and protein levels were measured via qRT–PCR and Western blotting. Cell viability was assessed by CCK-8 assay. The target relationship between SPI1 and hexokinase 2 (HK2) was determined using dual-luciferase reporter detection. ChIP was conducted to confirm the targeted relationship between SPI1 and the HK2 promoter. Immunohistochemistry analysis was conducted to measure the positive cell number of SPI1 and HK2 in melanoma tissues. The cell migration abilities were determined using a wound healing assay. Glucose consumption, pyruvate dehydrogenase activity, lactate production and ATP levels were measured to assess glycolysis. SPI1 transcription in melanoma cells and tissues was dramatically higher than that in adjacent normal tissues and epidermal melanocyte HEMa-LP, respectively. Knockdown of SPI1 restrained cell viability, metastasis and glycolysis in melanoma cells. SPI1 directly targeted HK2, and knockdown of SPI1 repressed HK2 expression. Overexpression of HK2 weakened the inhibitory effects of SPI1 knockdown on the viability, metastasis and glycolysis of melanoma cells. The serine–threonine kinase 1 (AKT1)/mammalian target of rapamycin (mTOR) axis is involved in melanoma progression. SPI1 knockdown restrained melanoma cell proliferation, metastasis and glycolysis by regulating the AKT1/mTOR pathway.

更新日期：2023-08-04

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