MITF / TFE (CLEAR)
MITF, TFE3, TFEB and TFEC are bHLH-LZ factors that dimerize only with each other, because a three-residue insertion kinks their leucine zipper. They read a canonical E-box core with a preference for T and A flanks, giving the palindromic 10-mer GTCACGTGAC, known as the CLEAR box in lysosomal and autophagy promoters; at the weaker CATGTG form of the core the 5' T is not merely preferred but required, and that version is the melanocyte M-box (TCATGTGA) of the pigmentation genes. MITF is the lineage factor of melanocytes and is mutated in Waardenburg syndrome type 2A and predisposes to melanoma; TFEB and TFE3 run the lysosome-autophagy program downstream of mTORC1 and are recurrently rearranged in renal cell carcinoma. The four paralogues cannot be told apart from sequence.
The factors
MITF, TFE3, TFEB and TFEC form the MiT/TFE family of basic helix-loop-helix leucine zipper (bHLH-LZ) factors. MITF was identified from the mouse microphthalmia locus, where mutations cause loss of melanocytes, deafness and small eyes 8343963; the three TFE genes were then recognized as a discrete family that dimerizes internally - each member homodimerizes and heterodimerizes with the others, but not with MYC, MAX or USF 7958932. The structural basis is a three-residue insertion that kinks the leucine zipper, which excludes the other bHLH-ZIP proteins from the dimer interface and explains why MiT/TFE factors, despite reading the same DNA core, form a closed dimerization set 23207919. Expression is what separates them: MITF is restricted (melanocytes, mast cells, osteoclasts, retinal pigment epithelium) and has multiple isoforms driven from alternative promoters, TFE3 and TFEB are broadly expressed and act as the general lysosomal-autophagy arm, and TFEC is largely myeloid. All four are regulated post-translationally rather than only transcriptionally; TFEB in particular is phosphorylated by mTORC1 at the lysosomal surface and held in the cytoplasm until nutrient stress releases it 22692423. The atlas assigns 14 motifs here (4 JASPAR, 6 HOCOMOCO, 4 CIS-BP) across all four symbols, cohesion 0.94, with TFE3.H14CORE.0.PSM.A as the representative.
The motif
The core is the canonical E-box CACGTG, read as two CAC half-sites by the histidine-glutamate-arginine triad in each basic region, exactly as in MAX 23207919. What distinguishes MiT/TFE is the flanking base, but the requirement is specific to one form of the core. At a CATGTG E-box a 5' T is essential: MITF bound TCATGTGA and not the same E-box lacking the T, correctly predicting that the CATGTG box of the P gene would not be bound, and the E-boxes of the tyrosinase, TRP-1, TRP-2 and QNR-71 promoters carry the T without exception 9819381. That T-flanked CATGTG is the classical melanocyte M-box 1321344,7958932. At a CACGTG E-box the same work saw no specific requirement for the flanking sequence 9819381, even though every MiT/TFE model in these databases is built with a T and the reciprocal A around the core, giving the palindromic 10-mer GTCACGTGAC. In the lysosomal literature that 10-mer is called the CLEAR element (Coordinated Lysosomal Expression And Regulation), GTCACGTGAC, found in the promoters of most lysosomal hydrolase, subunit and trafficking genes 19556463. So the M-box and the CLEAR box are one arrangement described in two fields, a T/A-flanked E-box, differing only in the central base pair of the core. Two consequences follow. First, MITF, TFE3, TFEB and TFEC are not separable from one another by sequence: pairwise similarity among their models in this atlas is 0.97-0.995. Second, the motif is essentially identical to the USF motif, and very close to BHLHE41 and CLOCK-BMAL1 models, so the T/A flanks buy discrimination against MYC-MAX but not against those. The family data also contains lower-information, ChIP-derived variants (a grade-C TFEB model reading gTCACATG) that lose the 3' half entirely.
Where it acts
MiT/TFE sites are promoter-proximal and CpG-island-associated. The CLEAR network was defined as a few hundred coordinately regulated lysosomal and autophagy genes carrying the element within the first few hundred base pairs of the transcription start site 19556463,21752829, and TFEB ChIP-seq recovers that set plus autophagosome-formation genes 21752829,21617040. MITF occupancy is layered on top of a melanocyte-specific accessibility landscape: the same GTCACGTGAC sequence is used at pigmentation genes (TYR, DCT, TYRP1, PMEL), at lysosome-related genes in melanosome biogenesis, and at proliferation genes, and MITF co-occupies melanocyte enhancers with SOX10 and the AP-1 machinery. TFEB and TFE3 occupancy is switched by subcellular localization rather than by motif availability - the sites are present and often accessible even when the factor is cytoplasmic 19556463,22692423.
Biology and disease
MITF is the lineage-determining factor of the melanocyte: loss-of-function alleles cause the mouse microphthalmia phenotypes and, in humans, Waardenburg syndrome type 2A (deafness with pigmentary anomalies) 8343963,7874167 and Tietz syndrome. The recurrent MITF E318K substitution, which impairs SUMOylation, raises melanoma risk and also associates with renal cell carcinoma 22080950,22012259. TFEB and TFE3 drive lysosomal biogenesis and autophagy downstream of mTORC1 and starvation 21617040,22692423, and are involved in lysosomal storage disease biology as therapeutic targets. Translocations involving these genes are diagnostic in cancer: promoter substitution at t(6;11)(p21;q13) drives TFEB overexpression in a distinct renal cell carcinoma subtype, and TFE3 fusions define another renal carcinoma group 12837690 and alveolar soft part sarcoma.
Reading a hit
A GTCACGTGAC or TCATGTGA hit identifies a T/A-flanked E-box. That is a real narrowing - it argues against MYC-MAX - but it does not name the factor: USF1/USF2, BHLHE41, CLOCK-BMAL1 and all four MiT/TFE proteins share it. Within the family, a hit cannot distinguish MITF from TFE3, TFEB or TFEC; the usual discriminator is expression, so a hit in melanocyte data suggests MITF and a hit in a starvation experiment suggests TFEB, and neither inference comes from the sequence. ChIP-derived MiT/TFE models are additionally biased toward active CpG-island promoters. Prefer the HOCOMOCO models with in vitro (S or M) support.
How this was worked out
Mutagenesis of the TRP-1 promoter identifies the M box, an 11-bp element conserved between the melanocyte-specific TRP-1 and tyrosinase promoters, as a positive regulatory element.
Still open then: The M-box-binding factor was not identified.
The mouse microphthalmia locus, whose alleles affect pigmentation, eye size, bone resorption, mast cells and hearing, is shown to encode a bHLH-ZIP protein.
MITF binds DNA as a homodimer or as a heterodimer with TFEB, TFE3 or TFEC, defining the MiT family, and activates pigmentation genes through the M box.
Still open then: Why MiT proteins do not partner other bHLH-ZIP factors.
A T immediately 5' of a CATGTG E-box is essential for MITF binding in vitro and in vivo, while no flank requirement was seen for CACGTG; pigmentation-gene E-boxes carry the T without exception.
Still open then: Later database models show a T/A flank around CACGTG as well, which this work did not detect.
Most lysosomal genes share a promoter-proximal GTCACGTGAC element (CLEAR) and are coordinately regulated by TFEB, which translocates to the nucleus under lysosomal storage stress.
- 2011highWhole-genome sequencing, linkage, case-control association, functional assays22080950 22012259
A germline MITF E318K substitution that impairs SUMOylation is associated with melanoma and with renal cell carcinoma.
- 2012highPhosphosite mapping and mutagenesis, kinase inhibition, localization imaging21617040 22692423
TFEB localization is set by phosphorylation, by ERK2 in response to nutrients and by lysosomal mTORC1 at Ser211 with 14-3-3 retention, so CLEAR occupancy is switched by nuclear entry rather than by site availability.
MITF crystal structures show a three-residue shift in the leucine zipper that kinks one helix and restricts heterodimerization, and an Ile212 nonpolar contact that explains M-box binding.
Still unresolved
- Whether the T and A flanks that every MiT/TFE model carries around CACGTG are a real binding preference or a property of the sequences the models were built from is untested: gel-shift and in vivo assays found the 5' T essential at CATGTG but no flanking requirement at CACGTG.
- Whether the M-box is more MiT/TFE-selective than the CLEAR box is not settled: the MITF structure attributes M-box binding to an Ile212 contact that related factors lack and reports that several of them bind the M-box weakly, but the JASPAR USF1 model also reads a T-flanked CATGTG (GTCATGTGAC).
- Whether MITF isoforms differ in DNA recognition is unclear: an alternatively spliced exon outside the bHLH-ZIP modulated DNA binding in vitro, but none of the database models resolves isoforms.
The T/A flanks make this motif distinguishable from MYC-MAX but not from USF1/USF2, BHLHE41 or CLOCK-BMAL1, which prefer the same extended palindrome; a GTCACGTGAC hit therefore names a group, not a protein. Within the family MITF, TFE3, TFEB and TFEC share one motif at 0.97-0.995 similarity, so assignment relies entirely on which factor is expressed and, for TFEB/TFE3, on whether it is nuclear at the time. Several models here are ChIP-only and low grade (a grade-C TFEB model reduces to gTCACATG), so they encode promoter context as much as intrinsic specificity. Prefer HT-SELEX or methyl-HT-SELEX-supported models when the question is direct binding.
References
- Hodgkinson CA, Moore KJ, Nakayama A, et al. Mutations at the mouse microphthalmia locus are associated with defects in a gene encoding a novel basic-helix-loop-helix-zipper protein. Cell 1993;74:395-404. PMID 8343963
- Hemesath TJ, Steingrímsson E, McGill G, et al. microphthalmia, a critical factor in melanocyte development, defines a discrete transcription factor family. Genes Dev 1994;8:2770-80. PMID 7958932
- Lowings P, Yavuzer U, Goding CR. Positive and negative elements regulate a melanocyte-specific promoter. Mol Cell Biol 1992;12:3653-62. PMID 1321344
- Aksan I, Goding CR. Targeting the microphthalmia basic helix-loop-helix-leucine zipper transcription factor to a subset of E-box elements in vitro and in vivo. Mol Cell Biol 1998;18:6930-8. PMID 9819381
- Pogenberg V, Ogmundsdóttir MH, Bergsteinsdóttir K, et al. Restricted leucine zipper dimerization and specificity of DNA recognition of the melanocyte master regulator MITF. Genes Dev 2012;26:2647-58. PMID 23207919
- Sardiello M, Palmieri M, di Ronza A, et al. A gene network regulating lysosomal biogenesis and function. Science 2009;325:473-7. PMID 19556463
- Palmieri M, Impey S, Kang H, et al. Characterization of the CLEAR network reveals an integrated control of cellular clearance pathways. Hum Mol Genet 2011;20:3852-66. PMID 21752829
- Settembre C, Di Malta C, Polito VA, et al. TFEB links autophagy to lysosomal biogenesis. Science 2011;332:1429-33. PMID 21617040
- Roczniak-Ferguson A, Petit CS, Froehlich F, et al. The transcription factor TFEB links mTORC1 signaling to transcriptional control of lysosome homeostasis. Sci Signal 2012;5:ra42. PMID 22692423
- Tassabehji M, Newton VE, Read AP. Waardenburg syndrome type 2 caused by mutations in the human microphthalmia (MITF) gene. Nat Genet 1994;8:251-5. PMID 7874167
- Yokoyama S, Woods SL, Boyle GM, et al. A novel recurrent mutation in MITF predisposes to familial and sporadic melanoma. Nature 2011;480:99-103. PMID 22080950
- Kuiper RP, Schepens M, Thijssen J, et al. Upregulation of the transcription factor TFEB in t(6;11)(p21;q13)-positive renal cell carcinomas due to promoter substitution. Hum Mol Genet 2003;12:1661-9. PMID 12837690
- Bertolotto C, Lesueur F, Giuliano S, et al. A SUMOylation-defective MITF germline mutation predisposes to melanoma and renal carcinoma. Nature 2011;480:94-8. PMID 22012259
14 motifs in this family — JASPAR 4 · HOCOMOCO 6 · CIS-BP 4
| motif | database | consensus | bits |
|---|---|---|---|
| TFEC.H14CORE.1.SM.BSMB | HOCOMOCO | gGTCACGTGAt | 17.9 |
| TFEC.H14CORE.0.M.BMB | HOCOMOCO | GTCACGTGAt | 16.7 |
| TFE3.H14CORE.0.PSM.APSMA | HOCOMOCO | gGTCACGTGAt | 15.8 |
| TFE3 (MA0831.3) | JASPAR | GTCACGTGAC | 15.4 |
| TFEB.H14CORE.0.SM.BSMB | HOCOMOCO | GTCACGTGAt | 15 |
| M04078_2.00 | CIS-BP | CACGTGAC | 14.5 |
| M08058_2.00 | CIS-BP | GTCACGTGAC | 14.5 |
| TFEC (MA0871.3) | JASPAR | CACGTGAC | 14.3 |
| M02727_2.00 | CIS-BP | gTCACGTGAc | 13.5 |
| MITF (MA0620.4) | JASPAR | gTCACGTGAc | 12.7 |
| MITF.H14CORE.0.P.BPB | HOCOMOCO | CaCGTGAC | 11.2 |
| TFEB (MA0692.2) | JASPAR | CACGTGAC | 10.8 |
| TFEB.H14CORE.1.P.CPC | HOCOMOCO | gTCACATG | 10.7 |
| M02773_2.00 | CIS-BP | CACGTGA | 9.4 |
HOCOMOCO tags read as assay + quality grade: P ChIP-seq, S HT-SELEX, M methyl-HT-SELEX, G GHT-SELEX, I SMiLE-seq, B PBM; grade A means two or more assay types agreed. A P-only matrix is a record of where the protein was crosslinked, which need not be a sequence it binds itself.
The factors, elsewhere
These entries are written from the primary literature and every citation is checked against PubMed, but this one can still be out of date, too confident, or simply wrong — especially where a motif’s published site and its database matrices disagree. Corrections and additions are welcome, and they are the fastest way to make this page better for everyone.