Shinde Anjali, Jung Hyeryeon, Lee Hayun, Singh Kritarth, Roy Milton, Gohel Dhruv, Kim Han Byeol, Mane Minal, Vasiyani Hitesh, Currim Fatema, Seo Yu Ri, Yang Seojin, Cho Ara, Yi Eugene C, Singh Rajesh
Department of Bio-Chemistry, Faculty of Science, The Maharaja Sayajirao University of Baroda, Sayajigunj, Vadodara, Gujarat, 390002, India.
Department of Molecular Medicine and Biopharmaceutical Sciences, Graduate School of Convergence Science and Technology, Seoul National University, Seoul, 03080, South Korea.
Cancer Metab. 2021 Apr 29;9(1):19. doi: 10.1186/s40170-021-00254-9.
Tumor necrosis factor-α (TNF-α) is an immunostimulatory cytokine that is consistently high in the breast tumor microenvironment (TME); however, its differential role in mitochondrial functions and cell survival in ER/PR +ve and ER/PR -ve breast cancer cells is not well understood.
In the current study, we investigated TNF-α modulated mitochondrial proteome using high-resolution mass spectrometry and identified the differentially expressed proteins in two different breast cancer cell lines, ER/PR positive cell line; luminal, MCF-7 and ER/PR negative cell line; basal-like, MDA-MB-231 and explored its implication in regulating the tumorigenic potential of breast cancer cells. We also compared the activity of mitochondrial complexes, ATP, and ROS levels between MCF-7 and MDA-MB-231 in the presence of TNF-α. We used Tumor Immune Estimation Resource (TIMER) webserver to analyze the correlation between TNF-α and mitochondrial proteins in basal and luminal breast cancer patients. Kaplan-Meier method was used to analyze the correlation between mitochondrial protein expression and survival of breast cancer patients.
The proteome analysis revealed that TNF-α differentially altered the level of critical proteins of mitochondrial respiratory chain complexes both in MCF-7 and MDA-MB-231, which correlated with differential assembly and activity of mitochondrial ETC complexes. The inhibition of the glycolytic pathway in the presence of TNF-α showed that glycolysis is indispensable for the proliferation and clonogenic ability of MDA-MB-231 cells (ER/PR -ve) as compared to MCF-7 cells (ER/PR +ve). The TIMER database showed a negative correlation between the expressions of TNF-α and key regulators of mitochondrial OXPHOS complexes in basal breast vs lobular carcinoma. Conversely, patient survival analysis showed an improved relapse-free survival with increased expression of identified proteins of ETC complexes and survival of the breast cancer patients.
The evidence presented in our study convincingly demonstrates that TNF-α regulates the survival and proliferation of aggressive tumor cells by modulating the levels of critical assembly factors and subunits involved in mitochondrial respiratory chain supercomplexes organization and function. This favors the rewiring of mitochondrial metabolism towards anaplerosis to support the survival and proliferation of breast cancer cells. Collectively, the results strongly suggest that TNF-α differentially regulates metabolic adaptation in ER/PR +ve (MCF-7) and ER/PR -ve (MDA-MB-231) cells by modulating the mitochondrial supercomplex assembly and activity.
肿瘤坏死因子-α（TNF-α）是一种免疫刺激细胞因子，在乳腺肿瘤微环境（TME）中一直处于高水平；然而，其在雌激素受体/孕激素受体（ER/PR）阳性和ER/PR阴性乳腺癌细胞中线粒体功能和细胞存活方面的差异作用尚不清楚。
在本研究中，我们使用高分辨率质谱法研究了TNF-α调节的线粒体蛋白质组，并在两种不同的乳腺癌细胞系中鉴定了差异表达的蛋白质，即ER/PR阳性细胞系；管腔型，MCF-7和ER/PR阴性细胞系；基底样，MDA-MB-231，并探讨了其在调节乳腺癌细胞致瘤潜力中的作用。我们还比较了在TNF-α存在下MCF-7和MDA-MB-231之间线粒体复合物的活性、ATP和活性氧水平。我们使用肿瘤免疫评估资源（TIMER）网络服务器分析基底样和管腔型乳腺癌患者中TNF-α与线粒体蛋白之间的相关性。采用Kaplan-Meier方法分析线粒体蛋白表达与乳腺癌患者生存率之间的相关性。
蛋白质组分析显示，TNF-α在MCF-7和MDA-MB-231中均差异改变了线粒体呼吸链复合物关键蛋白的水平，这与线粒体电子传递链（ETC）复合物的差异组装和活性相关。在TNF-α存在下对糖酵解途径的抑制表明，与MCF-7细胞（ER/PR阳性）相比，糖酵解对于MDA-MB-231细胞（ER/PR阴性）的增殖和克隆形成能力是不可或缺的。TIMER数据库显示，基底样乳腺癌与小叶癌中TNF-α的表达与线粒体氧化磷酸化（OXPHOS）复合物的关键调节因子之间呈负相关。相反，患者生存分析显示，随着ETC复合物鉴定蛋白表达的增加，乳腺癌患者的无复发生存率提高。
我们研究中提供的证据令人信服地表明，TNF-α通过调节参与线粒体呼吸链超复合物组织和功能的关键组装因子和亚基的水平来调节侵袭性肿瘤细胞的存活和增殖。这有利于将线粒体代谢重新连接到回补反应以支持乳腺癌细胞的存活和增殖。总体而言，结果强烈表明，TNF-α通过调节线粒体超复合物的组装和活性来差异调节ER/PR阳性（MCF-7）和ER/PR阴性（MDA-MB-231）细胞中的代谢适应。
