Abstract P3-05-01: Increased glycolysis directly activates transcriptional coactivator SRC-3 to promote aggressive metastatic breast cancer

Abstract

Abstract Reprogramming of metabolic pathways is one of the central hallmarks of cancer cell growth and survival to support anabolic and energetic demands. Rapidly proliferating tumor cells utilize increased amounts of glucose for energy generation and macromolecule biosynthesis, yet it is still poorly understood how increased glucose-flux accentuates aggressive metastatic tumor. Understanding the molecular and genetic regulators of these metabolic-programs is critical to discover new drug targets for cancer therapy. Here, we report that the oncogenic transcriptional coregulator steroid receptor coactivator-3 (SRC-3) is a critical sensor of increased tumor glycolysis and re-programs transcriptional responses to support breast cancer growth and metastasis. From an unbiased kinome-wide RNAi library screen, we identified the glycolytic enzyme PFKFB4 as a direct stimulator of SRC-3 activity. PFKFB4 is a bifunctional metabolic enzyme that contains both 'kinase' and 'phosphatase' domain, yet the functional role of these regulatory domains in tumor metabolism is not clear. Glucose stimulation promotes PFKFB4-dependent phosphorylation of SRC-3 (pSRC-3) which then enhances transcriptional coactivation by this coregulator. Mechanistic studies revealed that SRC-3 and PFKFB4 mutually cooperate to maintain high levels of purine biosynthesis by transcriptionally upregulating rate limiting enzymes transketolase (TKT), adenosine monophosphate deaminase-1 (AMPD1) and xanthine dehydrogenase (XDH). Knockdown of PFKFB4 or SRC-3 significantly decreased the expression of these enzymes resulting in reduced proliferation of breast tumor cells, and this functional deficiency could be rescued by exogenous addition of purines. Chromatin immunoprecipitation (ChIP) studies identified increased occupancy of pSRC-3 on target gene promoters, which overlapped with activating transcription factor 4 (ATF4) binding sites in the proximal promoter region. Immunoprecipitation of ATF4 identified increased interaction with pSRC-3 in glycolytic breast tumor cells, whereas knockdown of PFKFB4 or expression of a phospho-deficient SRC-3 mutant significantly reduced the SRC-3/ATF4 association. These findings suggest that upon enhanced glycolysis, PFKFB4 induced activation of SRC-3 by phosphorylation promotes expression of purine biosynthetic genes primarily by coactivating ATF4. Importantly, ablation of either SRC-3 or PFKFB4 suppresses in vivo breast tumor growth and metastasis, and these tumorigenic-effects cannot be simulated by a phospho-deficient SRC-3 mutant suggesting the PFKFB4-SRC-3 axis may be therapeutically exploited for treating breast cancer patients. [S.D. is supported by Susan G. Komen Award PDF14300468]. Citation Format: Dasgupta S, Zhu B, Nikolai BC, Putluri N, Foulds CE, Tsai SY, Tsai M-J, O'Malley BW. Increased glycolysis directly activates transcriptional coactivator SRC-3 to promote aggressive metastatic breast cancer [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P3-05-01.