The Mitf/TFE Family of Transcription Factors: Master Regulators of Organelle Signaling, Metabolism, and Stress Adaptation Logan Slade and Thomas Pulinilkunnil

Home , FNIP1, Folliculin, GPNMB, TFE3, TFEB, TFEC

Published OnlineFirst August 29, 2017; DOI: 10.1158/1541-7786.MCR-17-0320

Review Molecular Cancer Research The MiTF/TFE Family of Transcription Factors: Master Regulators of Organelle Signaling, Metabolism, and Stress Adaptation Logan Slade and Thomas Pulinilkunnil

Abstract

The microphthalmia family (MITF, TFEB, TFE3, and TFEC) of mote tumorigenesis is reviewed. Likewise, the emerging function transcription factors is emerging as global regulators of cancer cell of the Folliculin (FLCN) tumor suppressor in negatively regulat- survival and energy metabolism, both through the promotion of ing the MiT/TFE family and how loss of this pathway promotes lysosomal genes as well as newly characterized targets, such as cancer is examined. Recent reports are also presented that relate to oxidative metabolism and the oxidative stress response. In addi- the role of MiT/TFE–driven lysosomal biogenesis in sustaining tion, MiT/TFE factors can regulate lysosomal signaling, which cancer cell metabolism and signaling in nutrient-limiting condi- includes the mTORC1 and Wnt/b-catenin pathways, which are tions. Finally, a discussion is provided on the future directions and both substantial contributors to oncogenic signaling. This review unanswered questions in the ﬁeld. In summary, the research describes recent discoveries in MiT/TFE research and how they surrounding the MiT/TFE family indicates that these transcription impact multiple cancer subtypes. Furthermore, the literature factors are promising therapeutic targets and biomarkers for relating to TFE-fusion proteins in cancers and the potential cancers that thrive in stressful niches. Mol Cancer Res; 15(12); mechanisms through which these genomic rearrangements pro- 1637–43. 2017 AACR.

Introduction Lysosomal Expression and Regulation (CLEAR) elements are recognized by the MITF family, which in turn promotes gene MITF is an evolutionarily conserved transcription factor with transcription (9, 10). Genome-wide chromatin immunoprecip- homologs identified in C. elegans and Drosophila (1). The MITF itation sequencing analysis demonstrated direct binding of family encodes four distinct genes; MITF, TFEB, TFE3,and TFEB to CLEAR elements with concomitant increment in lyso- TFEC. Structurally, MITF genes constitute a double helix leucine somalproteins(11).GenesthataremostassociatedwithTFEB zipper motif, a transactivating zone, and a domain responsible regulation contain clusters of multiple CLEAR sequences. for DNA contact and binding. The MiT/TFE family of basic Genome-wide analysis for clustered CLEAR sequences identi- helix-loop-helix (bHLH) transcription factors recognizes the fied 471 direct TFEB targets, which include lysosomal acidifi- transcription initiation or E-box (Ephrussi boxes) sites cation and degradation enzymes along with autophagy, exo-, (CANNTG) in the genome (2). The initial identification of the endo-, and phagocytosis genes. Surprisingly, several gene tar- microphthalmia family of transcription factors revealed that gets of TFEB also included those executing glucose and lipid MiT/TFEs must homodimerize or heterodimerize with another metabolism, perhaps underscoring the direct connection member of the MiT/TFE family to activate transcription (3–6). between lysosomal function and metabolism (Fig. 1; ref. 11). Early studies into the function of these transcription factors Subsequent reports have also identified that both TFE3 and identified that mutations in MITF led to Waardenburg syn- MITF are capable of binding CLEAR sequence elements to drome type II, characterized by hypopigmentation and defects induce lysosomal biogenesis and autophagy in a comparable in ectodermal development (7), while murine homozygous manner (12, 13). TFEB knockouts fail to develop due to lack of placental vas- The autophagy–lysosome system is a catabolic cellular process cularization (8). More recently, MITF, TFEB, and TFE3 were for whole organelles, protein aggregates, and other macromole- identified as regulators of lysosomal function and metabolism. cules (14). During carcinogenesis, autophagy exerts an antitu- Numerous lysosomal and autophagy genes with one or more morigenic effect by degrading and/or recycling damaged cellular 10 base pair motifs (GTCACGTGAC) termed as Coordinated organelles, thereby blocking the accumulation of endogenous mutagens, and preventing further genomic alterations. However, following tumor induction, cancer cells coopt autophagy as a cell Department of Biochemistry and Molecular Biology, Faculty of Medicine, Dal- survival mechanism to promote nutrient reallocation for diverse housie University, Dalhousie Medicine New Brunswick, New Brunswick, Canada. cellular needs. Therefore, autophagy can suppress cancer devel- Corresponding Author: Thomas Pulinilkunnil, Department of Biochemistry and opment through its cytoprotective properties; however, once Molecular Biology, Dalhousie University, Dalhousie Medicine New Brunswick, cancer has developed, these same properties sustain survival of 100 Tucker Park Road, Saint John, New Brunswick E2L4L5, Canada. Phone: 1- the tumor (15). Cytoprotective and oncoprotective properties of 506-636-6973; Fax: 1-506-636-6001; E-mail: [email protected] autophagy include managing oxidative stress, preventing DNA doi: 10.1158/1541-7786.MCR-17-0320 damage, and supporting metabolism under nutrient-depleted 2017 American Association for Cancer Research. conditions (15, 16). Adaptive metabolic reprogramming of

www.aacrjournals.org 1637

Slade and Pulinilkunnil

Figure 1. Regulation and downstream targets of the Microphthalmia family of transcription factors: TFEB, TFE3, and MITF are negatively regulated through phosphorylation by mTORC1, where they are then restricted to the cytosol by chaperone protein 14-3-3. Phosphorylation is reversed by calcineurin (CaN), although no reports have yet investigated whether this is true for MITF, and the dephosphorylated proteins are free to enter the nucleus, where they bind to E-boxes, CLEAR sequences, and M-boxes to promote transcription of associated genes. Genes regulated by TFEB, TFE3, and MITF promote autophagy and lysosomal catabolism, along with mitochondrial biogenesis. Cell processes regulated by the microphthalmia TFs sustain cell metabolism through ensuring a supply of amino acids, which feed protein and nucleotide biosynthesis, while also regulating to supply of energy via mitochondrial oxidative phosphorylation. Gene targets downstream of CLEAR sequences also include those that respond to oxidative stress, such as key components of the glutathione system.

cancer cells provides them with the ability to utilize diverse fusion, places TFEB under the control of a more active promoter substrates as the building blocks for molecules necessary for resulting in a 60-fold higher expression (20). Crucially, the proliferation. Indeed, autophagy participates in this program resulting protein products from the gene fusion events still have through degradation of lipids and proteins within lysosomes functional basic helix-loop-helix domains and nuclear localiza- to derive substrates for nucleic acid and membrane biosynthe- tion signals, keeping the transcriptional activation function sis (14, 16). The microphthalmia family of transcription factors intact (18). It has been reported that lysosomal localization, are regulators of the autophagy–lysosome system (Fig. 1), and and thus inhibition, of MiT/TFEs requires the first 30 amino increasing evidence suggests that they also directly regulate acids, corresponding to exon 1 (21). All reported gene fusions metabolic and growth signaling pathways, and as such, re- eliminate exon 1 from the resulting protein, indicating that the present an attractive therapeutic target with wide potential fusion proteins are unlikely to be able to localize to the lyso- cancer. This review will discuss the prospect of MITF, TFEB, and some, suggesting a mechanism of constitutive activation (18). TFE3 as necessary elements for the viability of several cancer Further associating MiT/TFEs in renal neoplasia is a kidney- types with specific emphasis on changes in cellular autophagy specific TFEB overexpression mouse model that developed severe and metabolism. kidney enlargement with multiple cysts at 30 days following birth, while Ki-67–positive neoplastic lesions were detected as soon as 12 days after birth (22). Renal-specific TFEB overexpres- Role of TFEB and TFE3 as Oncogenes sion also resulted in liver metastasis in 23% of mice (22). There A number of studies have determined TFE3 and TFEB as being are no reports about the activity of TFE fusion proteins; however, oncogenes. Chromosomal translocations resulting in gene a case study has identified strong nuclear staining of the TFE3 fusions involving TFE3 or TFEB are implicated in the develop- fusion protein, and a cell line and xenograft model generated ment of sporadic renal cell carcinomas (RCC) and soft tissue from the patient maintained this nuclear localization (23), a sarcomas. These genetic rearrangements cause overexpression of result that has been reproduced in several other IHC screens of the TFE proteins (17–19), and in the case of TFEB-MALAT1 TFE3 and TFEB translocation cancers (17, 24, 25).

1638 Mol Cancer Res; 15(12) December 2017 Molecular Cancer Research

MIT/TFE Regulation of Lysosomal Autophagy

The role of autophagy in TFE fusion cancers remains con- D–CDK4/6 complex formation before becoming inhibitory to troversial. In the aforementioned mouse model of renal TFEB cell-cycle progression through CDK4 phosphorylation (35). overexpression, LC3 expression was unchanged when com- Interestingly, renal-specific TFEB overexpression in mice also pared with control mice, and crossing TFEB overexpression results in elevated cyclin D1 and p21 gene expression (22). mice with Atg7 knockout mice did not significantly reduce cancer development (22). Conversely, several reports have Upstream Regulators Modifying Oncogenic identified that cathepsin K immunoreactivity and expression – Outcomes of MiT/TFEs: Role of Lysosomal is a distinguishing feature of these neoplasms (26 28). Cathep- b sin K is a lysosomal cysteine protease, which is regulated by Signaling and Wnt/ -Catenin Pathways MITF in macrophages and osteoclasts (29), however, unlike FLCN–TFE axis cathepsins A, B, D, and F, does not contain and upstream Mutations in the FLCN gene result in Birt–Hogg–Dube (BHD) CLEAR sequence promoter (11). Despite lacking a CLEAR syndrome characterized by renal and pulmonary cysts, noncan- promoter, the CTSK gene is highly enriched along with other cerous tumors of the hair follicles, and an increased risk of RCC lysosomal genes in pancreatic cancers driven by autophagy (36). FLCN is proposed to act as a tumor suppressor through (30), and thus likely indicates a probable autophagy gene positive regulation of AMPK (AMP activated kinase; refs. 36, 37), signature in TFE translocation–driven RCCs. Given that RCCs and thus negative regulation of mTOR (38), which is supported by are characterized by metabolic dysregulation (31, 32), it is a homozygous knockout mouse model of BHD, which displayed tempting to speculate that TFE fusion proteins promote stress hyperactivation of mTOR (39). The FLCN tumor suppressor's response programs and help renal neoplasms to overcome relationship with mTOR is uncertain, given that reports describe metabolic crisis. There is preliminary support for this idea, as FLCN as a GAP (GTPase-activating protein) for Rag C/D (40). highlighted by increases in mTORC1 activity, the master reg- Given that GDP-loaded Rag C/D is necessary for amino acid ulator of growth and metabolism. TFE fusion RCCs display sensing by mTORC1, a GAP for these proteins will activate this elevated ribosomal S6 phosphorylation, a positive indicator of pathway (Fig. 2; refs. 41, 42). Indeed, models of BHD in yeast, mTORC1 activity, and therefore linking the MiT/TFE family of mammalian cancer cell lines, and mice show that FLCN knock- proteins with sustaining oncogenic anabolic pathways (33). down or heterozygous knockout results in reduced mTORC1 Molecular analysis of TFE fusion cancers also revealed elevated activity as measured by phosphorylation of S6 or S6K, while still expression of cell cycle–related proteins Cyclin D1 and D3 resulting in renal tumorigenesis (43–45). As FLCN seems to have along with p21 (CDKN1A; ref. 34), which promotes Cyclin conflicting roles in regulating mTORC1 and AMPK, it seems that

Figure 2. The microphthalmia transcription factors regulate signaling networks central to cancer: TFEB, TFE3, and MITF positively regulate genes promoting mTORC1 signaling (highlighted in red and italicized). Vacuolor ATPase (vATPase) activates Ragulator, a GEF for Rag A/B in the presence of amino acids, while the folliculin complex (FLCN, folliculin-interacting protein) acts as a GAP for Rag C/D. GTP-loaded Rag A/B and GDP-loaded Rag C/D recruit mTORC1 to the lysosome, where it can be activated. Activated Rag GTPases also recruit TFEB to the lysosome in combination with FLCN, where it is phosphorylated and inactivated by TFEB; however, in cancer, loss of inhibitory phosphorylation leads to an apparent feed-forward process, increasing mTOR activity. MITF and TFEB also participate in b-catenin activation through sequestration of the destruction complex into multivesicular bodies and subsequent degradation in lysosome. Degradation of the b-catenin destruction complex causes increased b-catenin activity, leading to cell proliferation.

www.aacrjournals.org Mol Cancer Res; 15(12) December 2017 1639

Slade and Pulinilkunnil

there are other mechanisms through which FLCN acts as tumor (55). Furthermore, constitutively active BRAF melanomas have suppressor and one candidate is through cytoplasmic sequestra- suppressed oxidative metabolism caused by downregulation of tion of MiT/TFE proteins. A report published in 2010 first the MITF–PGC1a axis. Conversely, therapeutic BRAF inhibi- highlighted that FLCN and TFE3 have a direct regulatory inter- tionresultsinanincreaseofMITF–PCG1a axis along with action in RCC (46). FLCN-null cells were shown to have decreased oxidative metabolism. The induction of oxidative metabolism TFE3 phosphorylation, which resulted in increased nuclear local- in MITF-overexpressing melanoma cells results in increased ization. FLCN-deficient cells also displayed greater TFE3 M-Box sensitivity to the mitochondrial uncoupler 2,4-dintrophenol, promoter activity and had elevated expression of MiT/TFE target revealing a novel biomarker for efficacy of antimetabolism genes, including several related to lysosomal activity, both in vitro therapeutics (56). and in BHD patients (46). FLCN-null cells also expressed elevated mRNA levels of GPNMB (glycoprotein nmb), a marker of mel- Regulation of mTORC1 by MiT/TFE anoma, glioma, breast cancers, which was also upregulated in a The role of MiT/TFEs in FLCN tumor progression also high- renal-specific TFEB overexpression mouse model (22). Further lights their role in lysosomal signaling and nutrient sensing. evidence to support a role for TFEB in FLCN tumor suppression Indeed, it is now understood that MITF and TFEB must be was published in 2013, wherein the authors showed that FLCN recruited to lysosomes to undergo inactivation by mTOR phos- loss led to increased nuclear TFEB caused by dysregulated lyso- phorylation, a process accomplished through GTP-loaded Rag somal signaling. The authors also confirmed that FLCN directly A/B, which have a direct interaction with MiT/TFEs (21). Recent interacts with Rag A/B in the absence of amino acids and promotes studies have identified the major players in TFEB regulation as GTP loading of Rag A/B, which is a prerequisite for mTOR mTORC1 (mTOR complex 1) and the ERK, which are central activation (45). The lysosomal surface is now understood to be regulators of anabolism and proliferation (57, 58). Phosphor- a center for nutrient sensing (47, 48), namely amino acids, ylation of TFEB by mTOR at serine 211 creates a 14-3-3 binding through a complex of proteins, including Rag GTPases, the site, which results in cytosolic retention of the transcription Ragulator complex, vATPase, as well as the folliculin complex, factor (57). In the absence of mTOR repression, TFEB is no containing both FLCN and FNIP1 (42). There is strong evidence to longer bound by 14-3-3 and is free to enter the nucleus where it indicate that FLCN is an activator of amino acid signaling to can enhance transcription of target genes (57). Subsequent mTORC1 through its GEF activity on Rag A/B and GAP activity on studies have concluded that both TFE3 and MITF are controlled Rag C/D. However, this role conflicts with the conventional through mTOR-mediated phosphorylation (Fig. 2; refs. 12, 30). wisdom that mTOR activity is required for tumor progression. This gives rise to a negative feedback loop, where activated Indeed, it seems there are few cases where mTORC1 inhibition can MiT/TFE promotes lysosomal biogenesis, and increases auto- promote cell growth, notably in nutrient-deplete conditions. Cell phagy, which in turn induces mTORC1 activation through cultures models of oncogenic transformation display increased increasing lysosomal amino acids, and transcriptional up- proliferation in the presence of the mTOR inhibitor Torin1 only regulation of mTOR signaling proteins, such as FNIP2, RagC/ when essential amino acids are absent, which is dependent on a D, and vATPase (11). Therefore, loss of inhibitory feedback by functional lysosome, while mouse models of pancreatic cancer mTORC1 on MiT/TFEs could promote oncogenic transforma- have a greater proliferative index in the interior, hypoxic tumor tion through constitutive mTOR activation and signaling regions after rapamycin treatment (49). Therefore, it is plausible (Fig. 2). Work published in 2015 supports this hypothesis, that FLCN loss causes an increased risk of neoplasia as cells where the authors confirmed that one of TFE3, MITF, or TFEB acquire the ability to cope with nutrient deprivation following was overexpressed in most human pancreatic ductal adenocar- constitutive activation of MiT/TFE proteins. In support of this cinoma (PDAC) cells and patient samples and showed consti- hypothesis are data that indicate that FLCN-null cells have greater tutive nuclear localization (30). Localization and activation of levels of autophagy proteins (50), while suppression of autop- MiT/TFEs in PDAC cells was not dependent on mTOR activation hagy in these cells results in increased sensitivity to paclitaxel or nutrient status, indicating a loss of inhibitory feedback. treatment (51). The constitutive activation and expression of MiT/TFEs resulted in elevated levels of autophagy–lysosome genes and increased FLCN and MiT/TFE in mitochondrial metabolism autophagic flux. Interestingly, mTOR activity remained constant FLCN loss also promotes significant metabolic remodeling, in PDAC cells even after 60 minutes of amino acid starvation, as indicated by an increase in mitochondrial biogenesis, which while siRNA knockdown of the overexpressed MiT/TFE rendered is dependent on PGC1a (52–54). Although metabolic changes mTOR amino acid sensitive. A metabolomics approach con- are thought to be a result of AMPK signaling (52, 53), it is likely firmed that levels of free amino acids were most affected by MiT/ that activation of MiT/TFE proteins in FLCN-deficient cells TFE knockdown, while overexpression of MITF in a noncancer- causes an upregulation of PCG1a, which is under the control ous pancreatic duct epithelial cells supported growth in amino of a CLEAR promoter, contributing to the phenotype. Like acid–deficient media. Further evidence for a MiT/TFE feed- cancers that have lost FLCN, a subset of human melanomas, forward mechanism in cancer was provided in a report pub- approximately 10%, are characterized by elevated PGC1a lished in 2017. The authors found that overexpression of MiT/ expression, which is driven by overexpression of MITF-M TFE genes in PDAC, RCC, and melanoma directly resulted in the (55). PGC1a is the master regulator of mitochondrial biogen- overexpression of RagD, which rendered mTORC1 insensitive to esis, and hence oxidative metabolism, so it is not surprising nutrient starvation, fueling cell proliferation and oncogenesis in that PGC1a-elevated tumors exhibit greater respiratory capac- an mTORC1-dependent manner (59). It is clear that MiT/TFEs ity and enhanced reactive oxygen species clearance. PGC1a- and autophagy can fuel cancer metabolism; however, there elevated tumors require PGC1a for proliferation, and gene remains several questions about the mechanisms that regulate knockdown of PGC1a renders the cells susceptible to apoptosis their expression and activity in cancer cells. It is currently

1640 Mol Cancer Res; 15(12) December 2017 Molecular Cancer Research

MIT/TFE Regulation of Lysosomal Autophagy

unknown as to why only one of the transcription factors (TFEB, Wnt signaling through similar mechanisms, as wild-type but TFE3, or MITF) is overexpressed in a particular cancer, and not AMPK DKO mouse embryos displayed extensive colocali- whether this has any relevance to the phenotype. Likewise, the zation between lysosomes and GSK3b (65). Wnt signaling and role of MITF in cancers beyond melanoma is understudied; gene expression is also upregulated in TFEB overexpression however, more recently, it has been shown in that MITF, but not renal cancer mouse models, while treating these mice with the splice variant MITF-M, is overexpressed in some pancreatic Wnt inhibitors successfully reduces tumor growth (22). cancers and thus should be researched for a role in tumorigen- Although the evidence is clear that MiT/TFEs feed into Wnt esis of other tissue types (30). Furthermore, questions remain signaling and vice versa (Fig. 2), the requirement of autophagy about how MiT/TFEs escape regulatory control by mTOR, given in the process is arguable. Indeed, autophagy can negatively that mTOR is commonly activated in many cancers. One pro- regulate b-catenin signaling upon nutrient deprivation through posed mechanism is through overexpression of nuclear impor- binding of LC3 to b-catenin and subsequent degradation (66). tin 8 (IPO8), which was identified as a common binding partner Likewise, autophagy can inhibit Wnt signal transduction of MiT/TFEs in PDAC cells, and knockdown of IPO8 decreased through degradation of dishevelled (DVL), the cytoplasmic nuclear localization of the transcription factors. It remains to be effector of Wnt receptor: frizzled (67). Although the association discovered exactly how IPO8 prevents mTOR inhibition; how- between the MiT/TFE family and Wnt signaling has only ever, there may be alternate mechanisms through which MiT/ recently become apparent, MITF has been linked to the Wnt TFEs become constitutively activated. Alterations in hetero- or pathwayinmelanomafarearlier.b-Catenin–induced melano- homodimerization frequency and spontaneity with other MiT/ ma growth requires functional MITF, and nuclear accumulation TFE family members, as well as incorrect spatial regulation, that of b-catenin was correlated with increased MITF expression is, failing to recruit MiT/TFEs to the lysosome where mTOR (68). Likewise, the melanocyte-specificisoform,MITF-M,is resides could also account for constitutive activity. directly regulated by downstream transcription factor of Wnt, LEF1, through binding of the upstream M promoter (69, 70). Wnt/b-catenin–TFE axis Conversely, MITF is regulated by noncanonical Wnt family A further mechanism through which MiT/TFEs can become members, namely WNT5A, which causes downregulation of implicated in cancer is through interplay between other known MITF and is characteristic of a distinct class of melanomas that oncogene networks, namely the Wnt/b-catenin pathway. The are resistant to BRAF inhibition and immunotherapy (71, 72). Wnt signaling pathway functions through promoting nuclear localization of b-catenin, an oncogenic transcription factor, as a Conclusion and Future Role of MiT/TFEs result of degrading the destruction complex, which includes Biology in Health and Disease GSK3b, AXIN, and APC among other proteins. b-Catenin is found to be constitutively activated in multiple cancer types as In conclusion, the microphthalmia family of transcription well as in mesenchymal and stem cell–like cancer cells (60–63). factors is emerging as important players in the development and Two reports from 2015 indicate that MiT/TFEs are under the sustainment of cancer. These transcription factors are activated direct regulation of Wnt signaling pathway member GSK3b as a or overexpressed in a diverse array of cancers where they are result of three conserved serine residues in the C-terminus involved in sustaining proliferation, driving metabolism, and region. Ploper and colleagues (13) showed that Wnt treatment overcoming stress. As a result, they represent an attractive ther- of melanoma cells results in increased MITF stability and apeutic target alone and in combination with other chemother- nuclear localization, while mutation of the putative GSK3b apeutic agents that induce cell stress. However, important ques- phosphorylation sites produced the same phenotype. With tions still need to be answered before this research can be regards to TFEB, another group (64) similarly noted that GSK3b translated to improved patient outcomes. A better understand- inhibition caused increased TFEB nuclear localization, as well ing of mechanisms surrounding activation in the cancer cell as lysosomal biogenesis and autophagy; however, the stability will help design therapies, especially as MiT/TFE inhibitors of the protein was unstudied. Interestingly, recent reports have tend to be oncogenic drivers. Furthermore, an understanding also highlighted the role of MiT/TFEs in promoting Wnt sig- of the systemic consequence of MiT/TFE inhibition must be naling through sequestration and degradation of the destruc- studied, given that these transcription factors are considered tion complex in autolysosomes. A tetracycline-inducible MITF essential for preventing neurodegeneration and cardiovascular melanoma cell line displayed greater Wnt reporter gene activity disease, and with respect to cancer, impacts on immune system following MITF induction, in a manner dependent on crucial function merit further investigation. endosome trafficking protein Vps27 (13). Furthermore, MITF Disclosure of Potential Conflicts of Interest induction in C32 melanoma cells caused colocalization of No potential conflicts of interest were disclosed. Axin1, the scaffold for the b-catenin destruction complex, with vesicular structures indicating that MITF induced sequestration Grant Support of the destruction complex as a mediator of Wnt signaling (13). ThisworkwasfundedbygrantsfromtheNaturalSciencesandEngi- Modulation of Wnt signaling through destruction complex neering Research Council of Canada (RGPIN-2014-03687) and Beatrice sequestration is not limited to MITF, as chronic TFEB inhibition Hunter Cancer Research Institute seed funding grant (to T. Pulinilkunnil). in AMPK double knockout (DKO) mouse embryos led to L. Slade is supported by the Cancer Research Training Program of the impaired endoderm differentiation due to increased b-catenin Beatrice Hunter Cancer Research Institute, with funds provided by the — phosphorylation resulting in decreased gene expression of Canadian Breast Cancer Foundation Atlantic Region and the New Bruns- wick Health Research Foundation. b-catenin targets. Wnt signaling was partially rescued in AMPK DKO mouse embryos through expression of constitutively Received June 22, 2017; revised August 8, 2017; accepted August 24, 2017; active TFEB. Interestingly, TFEB and MITF appear to mediate published OnlineFirst August 29, 2017.

www.aacrjournals.org Mol Cancer Res; 15(12) December 2017 1641

Slade and Pulinilkunnil

References 1. Hallsson JH, Haflidadottir BS, Stivers C, Odenwald W, Arnheiter H, 23. Hirobe M, Masumori N, Tanaka T, Kitamura H, Tsukamoto T. Establish- Pignoni F, et al. The basic helix-loop-helix leucine zipper transcription ment of an ASPL-TFE3 renal cell carcinoma cell line (S-TFE). Cancer Biol factor Mitf is conserved in Drosophila and functions in eye development. Ther 2013;14:502–10. Genetics 2004;167:233–41. 24. Altinok G, Kattar MM, Mohamed A, Poulik J, Grignon D, Rabah R. Pediatric 2. Betschinger J, Nichols J, Dietmann S, Corrin PD, Paddison PJ, Smith A. Exit renal carcinoma associated with Xp11.2 translocations/TFE3 gene fusions from pluripotency is gated by intracellular redistribution of the bHLH and clinicopathologic associations. Pediatr Dev Pathol 2005;8:168–80. transcription factor Tfe3. Cell 2013;153:335–47. 25. Argani P, Lae M, Hutchinson B, Reuter VE, Collins MH, Perentesis J, et al. 3. Fisher DE, Carr CS, Parent LA, Sharp PA. TFEB has DNA-binding and Renal carcinomas with the t(6;11)(p21;q12): clinicopathologic features oligomerization properties of a unique helix-loop-helix/leucine-zipper and demonstration of the specific alpha-TFEB gene fusion by immuno- family. Genes Dev 1991;5:2342–52. histochemistry, RT-PCR, and DNA PCR. Am J Surg Pathol 2005;29:230–40. 4. Hemesath TJ, Steingrímsson E, McGill G, Hansen MJ, Vaught J, 26. Martignoni G, Pea M, Gobbo S, Brunelli M, Bonetti F, Segala D, et al. Hodgkinson CA, et al. microphthalmia, a critical factor in melanocyte Cathepsin-K immunoreactivity distinguishes MiTF/TFE family renal trans- development, defines a discrete transcription factor family. Genes Dev location carcinomas from other renal carcinomas. Mod Pathol 2009; 1994;8:2770–80. 22:1016–22. 5. Muhle-Goll C, Gibson T, Schuck P, Schubert D, Nalis D, Nilges M, et al. The 27. Zheng G, Martignoni G, Antonescu C, Montgomery E, Eberhart C, Netto G, dimerization stability of the HLH-LZ transcription protein family is mod- et al. A broad survey of cathepsin K immunoreactivity in human neo- ulated by the leucine zippers: A CD and NMR study of TFEB and c-Myc. plasms. Am J Clin Pathol 2013;139:151–9. Biochemistry 1994;33:11296–306. 28. Lilleby W, Vlatkovic L, Meza-Zepeda LA, Revheim M-E, Hovig E. Transloca- 6. Pogenberg V, Ogmundsd€ ottir MH, Bergsteinsdottir K, Schepsky A, Phung B, tional renal cell carcinoma (t(6;11)(p21;q12) with transcription factor EB Deineko V, et al. Restricted leucine zipper dimerization and specificity of (TFEB) amplification and an integrated precision approach: a case report. DNA recognition of the melanocyte master regulator MITF. Genes Dev J Med Case Rep 2015;9:1–9. 2012;26:2647–58. 29. Motyckova G, Weilbaecher KN, Horstmann M, Rieman DJ, Fisher DZ, 7. Tassabehji M, Newton VE, Read AP. Waardenburg syndrome type 2 caused Fisher DE. Linking osteopetrosis and pycnodysostosis: regulation of by mutations in the human microphthalmia (MITF) gene. Nat Genet cathepsin K expression by the microphthalmia transcription factor family. 1994;8:251–5. Proc Natl Acad Sci 2001;98:5798–803. 8. Steingrimsson E, Tessarollo L, Reid SW, Jenkins NA, Copeland NG. The 30. Perera RM, Stoykova S, Nicolay BN, Ross KN, Fitamant J, Boukhali M, et al. bHLH-Zip transcription factor Tfeb is essential for placental vasculariza- Transcriptional control of autophagy-lysosome function drives pancreatic tion. Development 1998;125:4607. cancer metabolism. Nature 2015;524:361–5. 9. Sardiello M, Palmieri M, Ronza A, Medina DL, Valenza M, Gennarino VA. A 31. Hakimi AA, Reznik E, Lee CH, Creighton CJ, Brannon AR, Luna A, et al. An gene network regulating lysosomal biogenesis and function. Science integrated metabolic atlas of clear cell renal cell carcinoma. Cancer Cell 2009;325:473–7. 2016;29:104–16. 10. Martina JA, Diab HI, Li H, Puertollano R. Novel roles for the MiTF/TFE 32. Linehan WM, Srinivasan R, Schmidt LS. The genetic basis of kidney cancer: family of transcription factors in organelle biogenesis, nutrient sensing, a metabolic disease. Nat Rev Urol 2010;7:277–85. and energy homeostasis. Cell Mol Life Sci 2014;71:2483–97. 33. Argani P, Hicks J, De Marzo AM, Albadine R, Illei PB, Ladanyi M, et al. 11. Palmieri M, Impey S, Kang H, di Ronza A, Pelz C, Sardiello M, et al. Xp11 translocation renal cell carcinoma (RCC): extended immunohis- Characterization of the CLEAR network reveals an integrated control of tochemical profile emphasizing novel RCC markers. Am J Surg Pathol cellular clearance pathways. Hum Mol Genet 2011;20:3852–66. 2010;34:1295–303. 12. Martina JA, Diab HI, Lishu L, Jeong-A L, Patange S, Raben N, et al. The 34. Muller-H€ ocker€ J, Babaryka G, Schmid I, Jung A. Overexpression of cyclin nutrient-responsive transcription factor TFE3 promotes autophagy, lyso- D1, D3, and p21 in an infantile renal carcinoma with Xp11.2 TFE3-gene somal biogenesis, and clearance of cellular debris. Sci Signal 2014;7:ra9. fusion. Pathol Res Pract 2008;204:589–97. 13. Ploper D, Taelman VF, Robert L, Perez BS, Titz B, Chen HW, et al. MITF 35. Coleman ML, Marshall CJ, Olson MF. RAS and RHO GTPases in G1-phase drives endolysosomal biogenesis and potentiates Wnt signaling in mela- cell-cycle regulation. Nat Rev Mol Cell Biol 2004;5:355–66. noma cells. Proc Natl Acad Sci U S A 2015;112:E420–E9. 36. Schmidt LS, Linehan WM. Molecular genetics and clinical features of Birt- 14. Kaur J, Debnath J. Autophagy at the crossroads of catabolism and anab- Hogg-Dube syndrome. Nat Rev Urol 2015;12:558–69. olism. Nat Rev Mol Cell Biol 2015;16:461–72. 37. Baba M, Hong SB, Sharma N, Warren MB, Nickerson ML, Iwamatsu A, et al. 15. White E. Deconvoluting the context-dependent role for autophagy in Folliculin encoded by the BHD gene interacts with a binding protein, cancer. Nat Rev Cancer 2012;12:401–10. FNIP1, and AMPK, and is involved in AMPK and mTOR signaling. Proc 16. Kenific CM, Debnath J. Cellular and metabolic functions for autophagy in Natl Acad Sci 2006;103:15552–7. cancer cells. Trends Cell Biol 2015;25:37–45. 38. Shackelford DB, Shaw RJ. The LKB1-AMPK pathway: metabolism and 17. Argani P, Lal P, Hutchinson B, Lui MY, Reuter VE, Ladanyi M. Aberrant growth control in tumour suppression. Nat Rev Cancer 2009;9:563–75. nuclear immunoreactivity for TFE3 in neoplasms with TFE3 gene fusions: a 39. Hasumi Y, Baba M, Ajima R, Hasumi H, Valera VA, Klein ME, et al. sensitive and specific immunohistochemical assay. Am J Surg Pathol Homozygous loss of BHD causes early embryonic lethality and kidney 2003;27:750–61. tumor development with activation of mTORC1 and mTORC2. Proc Natl 18. Kauffman EC, Ricketts CJ, Rais-Bahrami S, Yang Y, Merino MJ, Bottaro DP, Acad Sci 2009;106:18722–7. et al. Molecular genetics and cellular features of TFE3 and TFEB fusion 40. Tsun ZY, Bar-Peled L, Chantranupong L, Zoncu R, Wang T, Kim C, et al. The kidney cancers. Nat Rev Urol 2014;11:465–75. folliculin tumor suppressor is a GAP for the RagC/D GTPases that signal 19. Klatte T, Streubel B, Wrba F, Remzi M, Krammer B, de Martino M, et al. amino acid levels to mTORC1. Mol Cell 2013;52:495–505. Renal cell carcinoma associated with transcription factor E3 expression and 41. Sancak Y, Peterson TR, Shaul YD, Lindquist RA, Thoreen CC, Bar-Peled L, Xp11.2 translocation. Am J Clin Pathol 2012;137:761–8. et al. The Rag GTPases bind raptor and mediate amino acid signaling to 20. Kuiper RP, Schepens M, Thijssen J, van Asseldonk M, van den Berg E, Bridge mTORC1. Science 2008;320:1496–501. J, et al. Upregulation of the transcription factor TFEB in t(6;11)(p21;q13)- 42. Kim J, Kim E. Rag GTPase in amino acid signaling. Amino Acids positive renal cell carcinomas due to promoter substitution. Hum Mol 2016;48:915–28. Genet 2003;12:1661–9. 43. van Slegtenhorst M, Khabibullin D, Hartman TR, Nicolas E, Kruger WD, 21. Martina JA, Puertollano R. Rag GTPases mediate amino acid–dependent Henske EP. The Birt-Hogg-Dube and tuberous sclerosis complex homologs recruitment of TFEB and MITF to lysosomes. J Cell Biol 2013;200: have opposing roles in amino acid homeostasis in schizosaccharomyces 475–91. pombe. J Biol Chem 2007;282:24583–90. 22. Calcagn A, Kors L, Verschuren E, De Cegli R, Zampelli N, Nusco E, et al. 44. Hartman TR, Nicolas E, Klein-Szanto A, Al-Saleem T, Cash TP, Simon MC, Modelling TFE renal cell carcinoma in mice reveals a critical role of WNT et al. The role of the Birt-Hogg-Dube protein in mTOR activation and renal signaling. eLife 2016;5:e17047. tumorigenesis. Oncogene 2009;28:1594–604.

1642 Mol Cancer Res; 15(12) December 2017 Molecular Cancer Research

MIT/TFE Regulation of Lysosomal Autophagy

45. Petit CS, Roczniak-Ferguson A, Ferguson SM. Recruitment of folliculin 59. Di Malta C, Siciliano D, Calcagni A, Monfregola J, Punzi S, Pastore N, et al. to lysosomes supports the amino acid-dependent activation of Rag Transcriptional activation of RagD GTPase controls mTORC1 and pro- GTPases. J Cell Biol 2013;202:1107–22. motes cancer growth. Science 2017;356:1188. 46. Hong SB, Oh H, Valera VA, Baba M, Schmidt LS, Linehan WM. Inac- 60. Cleary SP, Jeck WR, Zhao X, Chen K, Selitsky SR, Savich GL, et al. Iden- tivation of the FLCN tumor suppressor gene induces TFE3 transcrip- tification of driver genes in hepatocellular carcinoma by exome sequenc- tional activity by increasing its nuclear localization. PLoS One 2010; ing. Hepatology (Baltimore, Md) 2013;58:1693–702. 5:e15793. 61. Morin PJ, Sparks AB, Korinek V, Barker N, Clevers H, Vogelstein B, et al. 47. Settembre C, Fraldi A, Medina DL, Ballabio A. Signals from the lysosome: a Activation of b-catenin-Tcf signaling in colon cancer by mutations in control centre for cellular clearance and energy metabolism. Nat Rev Mol b-catenin or APC. Science 1997;275:1787. Cell Biol 2013;14:283–96. 62. Lamouille S, Xu J, Derynck R. Molecular mechanisms of epithelial– 48. Perera RM, Zoncu R. The lysosome as a regulatory hub. Annu Rev Cell Dev mesenchymal transition. Nat Rev Mol Cell Biol 2014;15:178–96. Biol 2016;32:223–53. 63. Vermeulen L, De Sousa E Melo F, van der Heijden M, Cameron K, de Jong 49. Palm W, Park Y, Wright K, Pavlova NN, Tuveson DA, Thompson CB. JH, Borovski T, et al. Wnt activity defines colon cancer stem cells and is The utilization of extracellular proteins as nutrients is suppressed by regulated by the microenvironment. Nat Cell Biol 2010;12:468–76. mTORC1. Cell 2015;162:259–70. 64. Marchand B, Arsenault D, Raymond-Fleury A, Boisvert FM, Boucher MJ. 50. Dunlop EA, Seifan S, Claessens T, Behrends C, Kamps MA, Rozycka E, Glycogen synthase kinase-3 (GSK3) inhibition induces prosurvival auto- et al. FLCN, a novel autophagy component, interacts with GABARAP phagic signals in human pancreatic cancer cells. J Biol Chem 2015;290: and is regulated by ULK1 phosphorylation. Autophagy 2014;10: 5592–605. 1749–60. 65. Young NP, Kamireddy A, Van Nostrand JL, Eichner LJ, Shokhirev MN, 51. Zhang Q, Si S, Schoen S, Chen J, Jin XB, Wu G. Suppression of autophagy Dayn Y, et al. AMPK governs lineage specification through Tfeb- enhances preferential toxicity of paclitaxel to folliculin-deficient renal dependent regulation of lysosomes. Genes Dev 2016;30:535–52. cancer cells. J Exp Clin Cancer Res 2013;32:99. 66. Petherick KJ, Williams AC, Lane JD, Ordonez-Mor~ an P, Huelsken J, 52. Yan M, Gingras MC, Dunlop EA, Nouet Y, Dupuy F, Jalali Z, et al. The tumor Collard TJ, et al. Autolysosomal b-catenin degradation regulates Wnt- suppressor folliculin regulates AMPK-dependent metabolic transforma- autophagy-p62 crosstalk. EMBO J 2013;32:1903. tion. J Clin Invest 2014;124:2640–50. 67. Gao C, Cao W, Bao L, Zuo W, Xie G, Cai T, et al. Autophagy negatively 53. Yan M, Audet-Walsh E, Manteghi S, Dufour CR, Walker B, Baba M, et al. regulates Wnt signalling by promoting Dishevelled degradation. Nat Cell Chronic AMPK activation via loss of FLCN induces functional beige Biol 2010;12:781–90. adipose tissue through PGC-1a/ERRa. Genes Dev 2016;30:1034–46. 68. Widlund HR, Horstmann MA, Price ER, Cui J, Lessnick SL, Wu M, et al. 54. Hasumi H, Baba M, Hasumi Y, Huang Y, Oh H, Hughes RM, et al. b-catenin–induced melanoma growth requires the downstream target Regulation of mitochondrial oxidative metabolism by tumor suppressor Microphthalmia-associated transcription factor. J Cell Biol 2002;158: FLCN. J Natl Cancer Inst 2012;104:1750–64. 1079–87. 55. Vazquez F, Lim JH, Chim H, Bhalla K, Girnun G, Pierce K, et al. PGC1a 69. Dorsky RI, Raible DW, Moon RT. Direct regulation of nacre, a zebrafish expression defines a subset of human melanoma tumors with increased MITF homolog required for pigment cell formation, by the Wnt pathway. mitochondrial capacity and resistance to oxidative stress. Cancer Cell Genes Dev 2000;14:158–62. 2013;23:287–301. 70. Saito H, Yasumoto KI, Takeda K, Takahashi K, Yamamoto H, Shibahara S. 56. Haq R, Shoag J, Andreu-Perez P, Yokoyama S, Edelman H, Rowe GC, et al. Microphthalmia-associated transcription factor in the Wnt signaling path- Oncogenic BRAF regulates oxidative metabolism via PGC1a and MITF. way. Pigment Cell Res 2003;16:261–5. Cancer Cell 2013;23:302–15. 71. Dissanayake SK, Olkhanud PB, O'Connell MP, Carter A, French AD, 57. Roczniak-Ferguson A, Petit CS, Froehlich F, Qian S, Ky J, Angarola B, et al. Camilli TC, et al. Wnt5A regulates expression of tumor associated antigens The transcription factor TFEB links mTORC1 signaling to transcriptional in melanoma via changes in STAT3 phosphorylation. Cancer Res control of lysosome homeostasis. Sci Signal 2012;5:ra42. 2008;68:10205–14. 58. Settembre C, Zoncu R, Medina DL, Vetrini F, Erdin S, Erdin S, et al. A 72. Wellbrock C, Arozarena I. Microphthalmia-associated transcription factor lysosometonucleus signalling mechanism senses and regulates the lyso- in melanoma development and MAP-kinase pathway targeted therapy. some via mTOR and TFEB. EMBO J 2012;31:1095. Pigment Cell Melanoma Res 2015;28:390–406.

www.aacrjournals.org Mol Cancer Res; 15(12) December 2017 1643

The MiTF/TFE Family of Transcription Factors: Master Regulators of Organelle Signaling, Metabolism, and Stress Adaptation

Logan Slade and Thomas Pulinilkunnil

Mol Cancer Res 2017;15:1637-1643. Published OnlineFirst August 29, 2017.

Updated version Access the most recent version of this article at: doi:10.1158/1541-7786.MCR-17-0320

Cited articles This article cites 72 articles, 26 of which you can access for free at: http://mcr.aacrjournals.org/content/15/12/1637.full#ref-list-1

Citing articles This article has been cited by 6 HighWire-hosted articles. Access the articles at: http://mcr.aacrjournals.org/content/15/12/1637.full#related-urls

E-mail alerts Sign up to receive free email-alerts related to this article or journal.

Reprints and To order reprints of this article or to subscribe to the journal, contact the AACR Publications Department at Subscriptions [email protected].

Permissions To request permission to re-use all or part of this article, use this link http://mcr.aacrjournals.org/content/15/12/1637. Click on "Request Permissions" which will take you to the Copyright Clearance Center's (CCC) Rightslink site.