Tfeb Functions As a Tumor Suppressor That Harnesses Myc-Induced Lymphomagenesis By Provoking Senescence and Metabolic Catastrophe

Metabolic reprogramming is a hallmark of cancer. MYC oncoproteins orchestrate the control of many aspects of metabolism, including mitochondrial biogenesis, glycolysis and glutaminolysis. This response allows the highly proliferative cancer cell to meet the increased demands for macromolecules and energy. Here we report that MYC promotes anabolism by actively suppressing the autophagy-lysosomal catabolic pathway. Notably, disabling this response by reactivating catabolism provokes metabolic catastrophe and senescence in MYC-driven lymphomas.

Gene expressing profiling of B220⁺ B cells from bone marrow of wild type (Wt) and pre-cancerous Eµ-Myc mice, and of Eµ-Myc lymphoma, demonstrated that genes involved in autophagy and lysosomal biogenesis are suppressed in MYC-expressing B cells. This response was due to MYC, as activation of the c-MYC transgene in human P493-6 B cells suppressed the expression of autophagy and lysosomal biogenesis genes. Further, this response was regulated by physiological cues that control endogenous c-Myc expression, as it was evident in naïve mouse splenic B cells treated with mitogen and in cytokine-treated hematopoietic cells. Finally, the suppression of autophagy-lysosomal genes was a hallmark of human malignancies with MYC involvement, including Burkitt lymphoma and colorectal cancer. Importantly, these changes in autophagy-lysosomal gene expression have functional consequences, as there are marked decreases in lysosomes in MYC-expressing versus control mouse embryonic fibroblasts and primary B cells, and in P493-6 B cells following MYC activation. Finally, consistent with an impaired autophagy-lysosomal pathway, there were marked increases in the levels of p62/Sequestrin, a receptor for ubiquitylated cargo on autophagosomes that is normally degraded by the autophagosomal pathway, in premalignant and neoplastic B220⁺ Eµ-Myc B cells.

A master regulator of autophagy and lysosomal biogenesis is TFEB that, like MYC, functions as a basic helix-loop-helix leucine zipper transcription factor. Our studies suggest that MYC can block TFEB function at three levels. First, TFEB expression is suppressed in human malignancies with MYC involvement, suggesting that in some contexts MYC may repress TFEB transcription. Second, MYC can repress TFEB transcription targets by competing with TFEB for binding to autophagy-lysosome gene targets. Specifically, the DNA recognition sequence of TFEB (CLEAR sequence) overlaps with the E-Box sequence of MYC, and chromatin immunoprecipitation analyses showed that MYC bound to these recognition elements in select TFEB targets and repressed their expression. Finally, MYC-expressing B cells display activation of mTORC1, which phosphorylates TFEB and blocks its nuclear localization. Specifically, MYC-expressing B cells have marked increases in the phosphorylation of the mTORC1 substrates S6K and 4EBP1, and show cytoplasmic localization of TFEB.

Given these findings, we hypothesized that MYC-transformed lymphoma cells might be sensitive to activating this catabolic pathway. We tested this hypothesis in Eµ-Myc;Rosa26-rtTa lymphoma cells that express the reverse tetracycline transactivator. These lymphoma cells were transduced with a pTight doxycycline- (Dox)-inducible retrovirus that expresses a constitutively active (mTORC1-resistant and nuclear) form of TFEB (TFEB-S211A) and the effects of the induction of TFEB were monitored. Importantly, our analyses established that the induction of TFEB-S211A markedly impaired the proliferation and tumorigenic potential of Eµ-Myc lymphoma. Mechanistically, this proliferative arrest was associated with the induction of the senescence program, and with an increased extracellular acidification rate. Strikingly, this metabolic shift was associated with reductions in oxygen consumption rate and reduced numbers of mitochondria. Therefore, TFEB functions as a tumor suppressor that harnesses MYC-induced tumorigenesis by inducing senescence and disabling mitochondrial anabolic pathways that are needed to sustain the growth of malignant cells.