Résumé
Background: The proto-oncogene MYC encodes a bHLHLZ transcription factor that binds to its obligate DNA-binding partner, MAX, leading to heterodimer occupancy of E-Box motifs within the genome. MYC is normally mitogen-dependent and tightly regulated where it plays a role in transcriptionally coordinating metabolic sufficiency for cell cycle progression, regulating both proliferation and viability. MYC is deregulated in the majority of human cancers. Importantly, MYC exists within an extended family of related bHLHLZ transcription factors including the MYC antagonizing MAX dimerization (MXD) proteins, MYC paralogs, such as MYCN, as well as MAX-like protein X (MLX), and its nutrient-sensing heterodimerization partners MondoA and carbohydrate response element binding protein (ChREBP). We recently defined a role for the extended MAX/MLX network in the regulation of metabolism and survival of cancer cells specifically with deregulated MYC or MYCN, in which MYC-MAX or MYCN-MAX heterodimers coordinately regulate transcription of important metabolic targets with MondoA-MLX to facilitate tumor cell survival. Targeting a number of these downstream effectors is sufficient to kill cancer cells with deregulated MYC. Methods: Expanding on our initial findings we have used molecular and biochemical approaches to demonstrate non-redundant roles for both MondoA-MLX and ChREBP-MLX in the transcriptional regulation of metabolism and survival of human cancer cells with deregulated MYC, including liver and colon cancer cells. We have also characterized the metabolic and biological phenotypes associated with targeted deletion of Mlx in the mouse, combining in vivo and ex vivo analysis with both whole-body deletion and tissue-specific ablation to unmask novel functions of Mlx and its binding partners. Results: Our results demonstrate that MLX and its binding partners regulate pathways associated with mitochondrial function as well as glucose and lipid metabolism in a non-redundant, cell-type-dependent manner. The presence of MLX stabilizes MondoA and ChREBP, and deletion of Mlx in the mouse leads to decreased levels of MondoA and Chrebp proteins in vivo. Whole-body deletion of Mlx in the mouse is well tolerated but leads to male infertility and teratozoospermia. This sterility phenotype is recapitulated by specific deletion of Mlx in the Sertoli cells of the testis, a major metabolic cell type within the tissue and the central regulator of multiple steps associated with germ cell fate, including production and/or transport of critical metabolites required by the germ cells. This indicates both cell-autonomous (germ cell) and non-cell-autonomous (Sertoli cell) regulation of survival and metabolism in spermatogenesis mediated by Mlx. Importantly, human patients with teratozoospermia exhibit diminished expression of MondoA, MLX and their metabolic targets. Moreover, overexpression of MYCN, MondoA, MLX and their metabolic targets occurs in patients with germ cell-derived tumors, such as seminomas. Conclusions: Transcriptional coordination between the mitogen- and nutrient-responsive arms of the MAX/MLX network exhibits context-dependent lethality. MondoA and Chrebp exhibit non-redundant functions in both cancer cells and normal mouse tissue, and inactivation of Mlx phenocopies loss of both factors, suggesting that targeting Mlx can inhibit both MondoA and Chrebp transcriptional activity and subsequent metabolic output. Metabolic regulation by Mlx impacts normal testicular tissue homeostasis and differentiation in the mouse, and its deregulation is associated with human diseases such as loss of function associated with infertility and gain of function associated with germ cell malignancy. While deletion of Myc, Mycn or Max is embryonic lethal in the mouse, deletion of MondoA, Chrebp or Mlx is well tolerated, indicating that targeting this arm of the extended MYC network, or their critical downstream targets, could be of therapeutic value in metabolic syndrome, fertility and cancer. Citation Format: Patrick A. Carroll, Daniel Diolaiti, Pei-Feng Cheng, Haiwei Gu, Danijel Djukovic, Daniel Raftery, Donald E. Ayer, Charles H. Muller, Robert N. Eisenman. Transcriptional regulation of metabolism by MLX and its binding partners is essential for tumor cell survival and spermatogenesis. [abstract]. In: Proceedings of the AACR Special Conference: Metabolism and Cancer; Jun 7-10, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(1_Suppl):Abstract nr PR12.