1. Academic Validation
  2. Low levels of pyruvate induced by a positive feedback loop protects cholangiocarcinoma cells from apoptosis

Low levels of pyruvate induced by a positive feedback loop protects cholangiocarcinoma cells from apoptosis

  • Cell Commun Signal. 2019 Mar 12;17(1):23. doi: 10.1186/s12964-019-0332-8.
Mingming Zhang 1 2 3 Yida Pan 4 Dehua Tang 1 Robert Gregory Dorfman 5 Lei Xu 6 Qian Zhou 3 Lixing Zhou 1 Yuming Wang 1 Yang Li 6 Yuyao Yin 1 Bo Kong 1 7 Helmut Friess 7 Shimin Zhao 2 3 Jian-Lin Wu 8 Lei Wang 9 Xiaoping Zou 10
Affiliations

Affiliations

  • 1 Department of Gastroenterology, Nanjing Drum Tower Hospital, the Affiliated Hospital of Nanjing University Medical School, Nanjing University, No.321 Zhongshan Road, 210008, Nanjing, People's Republic of China.
  • 2 Key laboratory of Reproduction Regulation of NPFPC (SIPPR, IRD), Fudan University, Shanghai, 200032, China.
  • 3 School of Life Sciences, Fudan University, Shanghai, China.
  • 4 Department of Digestive Diseases of Huashan Hospital, Shanghai, China.
  • 5 Northwestern University Feinberg School of Medicine, Chicago, IL, USA.
  • 6 Department of Gastroenterology, Nanjing Medical University Affiliated Drum Tower Clinical Medical College, Nanjing Medical University, Nanjing, China.
  • 7 Department of Surgery, Technical University of Munich (TUM), Munich, Germany.
  • 8 State Key Laboratory of Quality Research in Chinese Medicine, Macau Institute for Applied Research in Medicine and Health, Faculty of Chinese Medicine, Macau University of Science and Technology, Avenida Wai Long, Taipa, Macao, 442000, People's Republic of China. jlwu@must.edu.mo.
  • 9 Department of Gastroenterology, Nanjing Drum Tower Hospital, the Affiliated Hospital of Nanjing University Medical School, Nanjing University, No.321 Zhongshan Road, 210008, Nanjing, People's Republic of China. 867152094@qq.com.
  • 10 Department of Gastroenterology, Nanjing Drum Tower Hospital, the Affiliated Hospital of Nanjing University Medical School, Nanjing University, No.321 Zhongshan Road, 210008, Nanjing, People's Republic of China. zouxp@nju.edu.cn.
Abstract

Background: Cancer cells avidly consume glucose and convert it to lactate, resulting in a low pyruvate level. This phenomenon is known as the Warburg effect, and is important for cell proliferation. Although cMyc has often been described as an oncoprotein that preferentially contributes to the Warburg effect and tumor proliferation, mechanisms of action remain unclear. Histone deacetylase 3 (HDAC3) regulates gene expression by removing acetyl groups from lysine residues, as well as has an oncogenic role in Apoptosis and contributes to the proliferation of many Cancer cells including cholangiocarcinoma (CCA). HDAC inhibitors display antitumor activity in many Cancer cell lines. Cancer cells maintain low levels of pyruvate to prevent inhibition of HDAC but the mechanisms remain elusive. The purpose of our study was to explore the role of cMyc in regulating pyruvate metabolism, as well as to investigate whether the inhibitory effect of pyruvate on HDAC3 could hold promise in the treatment of Cancer cells.

Methods: We studied pyruvate levels in CCA cell lines using metabolite analysis, and analyzed the relationship of pyruvate levels and cell proliferation with cell viability analysis. We cultivated CCA cell lines with high or low levels of pyruvate, and then analyzed the protein levels of HDAC3 and apoptotic markers via Western Blotting. We then explored the reasons of low levels of pyruvate by using seahorse analysis and 13C6 metabolites tracing analysis, and then confirmed the results using patient tissue protein samples through Western Blotting. Bioinformatics analysis and transfection assay were used to confirm the upstream target of the low levels of pyruvate status in CCA. The regulation of cMyc by HDAC3 was studied through immunoprecipitation and Western Blotting.

Results: We confirmed downregulated pyruvate levels in CCA, and defined that high pyruvate levels correlated with reduced cell proliferation levels. Downregulated pyruvate levels decreased the inhibition to HDAC3 and consequently protected CCA cells from Apoptosis. Synergistically upregulated LDHA, PKM2 levels resulted in low levels of pyruvate, as well as poor patient survival. We also found that low levels of pyruvate contributed to proliferation of CCA cells and confirmed that the upstream target is cMyc. Conversely, high activity of HDAC3 stabilized cMyc protein by preferential deacetylating cMyc at K323 site, which further contributed to the low pyruvate levels. Finally, this creates a positive feedback loop that maintained the low levels of pyruvate and promoted CCA proliferation.

Conclusions: Collectively, our findings identify a role for promoting the low pyruvate levels regulated by c-Myc, and its dynamic acetylation in Cancer cell proliferation. These targets, as markers for predicting tumor proliferation in patients undergoing clinical treatments, could pave the way towards personalized therapies.

Keywords

Apoptosis; Cholangiocarcinoma; HDAC3; Pyruvate; cMyc.

Figures
Products
  • Cat. No.
    Product Name
    Description
    Target
    Research Area
  • HY-13909
    99.81%, HDAC3 抑制剂