|Coordinate||5,770,554 bp (GRCm38)|
|Base Change||T ⇒ A (forward strand)|
|Gene Name||zinc finger E-box binding homeobox 1|
|Synonym(s)||3110032K11Rik, Tw, MEB1, Zfhx1a, Zfhep, ZEB, AREB6, Zfx1a, Tcf18, Nil2, Tcf8, [delta]EF1|
|Chromosomal Location||5,591,860-5,775,467 bp (+)|
|MGI Phenotype||Mutations at this locus affect thymus organization and homozygotes exhibit severe thymic T cell deficiency. Some mutations also exhibit extensive skeletal abnormalities. Homozygotes generally die at birth.|
|Amino Acid Change||Tyrosine changed to Stop codon|
|Institutional Source||Beutler Lab|
Y902* in Ensembl: ENSMUSP00000025081 (fasta)
|Gene Model||not available|
|Phenotypic Category||T-dependent humoral response defect- decreased antibody response to rSFV, T-independent B cell response defect- decreased TNP-specific IgM to TNP-Ficoll immunization|
|Alleles Listed at MGI|
|Mode of Inheritance||Autosomal Recessive|
|Local Stock||Live Mice, Sperm, gDNA|
|Last Updated||05/13/2016 3:09 PM by Anne Murray|
|Record Created||11/10/2009 12:00 AM|
The cellophane phenotype was identified in the T-dependent humoral response screen and in the T-independent B cell response screen. Cellophane mice were unable to mount T-independent IgM responses following immunization with (4-hydroxy-3-nitrophenylacetyl)-Ficoll (NP-Ficoll) (Figure 1, left), and made suboptimal T-dependent IgG responses to a recombinant Semliki Forest virus (rSFV)-encoded β-galactosidase (rSFV-βGal) (Figure 1, right) (1). The frequency of mature B cells in the bone marrow and spleen of cellophane mice (Figure 2A and B) was normal, although B cells from cellophane mice expressed lower surface levels of IgD than B cells from wild type mice (1). However, cellophane mice had reduced frequencies of splenic marginal zone (MZ) B cells (Figure 2B) and peritoneal B1 B cells (Figure 2D); the frequencies of B1a and B1b B cells were equally affected (Figure 2E) (1). Cellophane mice had small, hypocellular thymi, and significant changes in thymic cell subsets (i.e. DN and SP thymocyte frequencies were increased; DP thymocyte frequencies were decreased) (Figure 2F) (1). The frequency of splenic T cells was not affected, but splenic NK cells were reduced (Figure 2H). Cellophane mice had normal levels of all Ig subtypes assayed with the exception of IgG2a, which was slightly reduced (1).
Seven days post-immunization with NP-CGG, cellophane mice had undetectable levels of NP-specific IgM (Figure 3A) and IgG1 (Figure 3B) (1). At day 14 post-immunization, the cellophane NP-specific IgG1 response was reduced compared to wild-type animals (Figure 3B), while NP-specific IgM remained undetectable (1). The IgG1 in the cellophane mice was determined to be low affinity at 28 days post-immunization (Figure 3C), indicating that these mice do not support functional germinal centers (GCs). The frequency of GC B cells was reduced in the cellophane mice (Figure 4A and C) (1). In addition, splenic B cells from immunized cellophane mice were unable to transfer NP-specific memory to naïve wild type mice (Figure 5).
|Nature of Mutation|
The cellophane mutation was mapped by bulk segregation analysis (BSA) of F2 intercross offspring using C57BL/10J as the mapping strain (n=18 with mutant phenotype, 82 with normal phenotype). The mutation showed strongest linkage with the marker at position 15408257 bp on Chromosome 18 (synthetic LOD=7.9) (Figure 6). Whole genome SOLiD sequencing identified a T to A transversion at base pair 5770552, 9.6 Mb from the lineage peak, on Chromosome 18 in the genomic region NC_000084. This position corresponds to base pair 2725 of the Zeb1 transcript, in exon 7 of 8 exons (Figure 7).
The mutated nucleotide is indicated in red lettering, and results in conversion of a tyrosine codon at position 902 to a premature stop codon.
Zeb1 and Zeb2 form the mammalian Zeb family of transcription factors. Their effects are typically repressive, although they have also been shown to activate the transcription of certain genes. Mouse Zeb1 and Zeb2 are 42% identical in sequence overall, and approximately 90% identical across their two zinc finger arrays (see below). Human and mouse Zeb1 are 88% identical.
Zeb1 contains seven C2H2-type zinc finger domains, each approximately 23 amino acids in length (2-5) (Figure 8). Four are clustered near the N-terminus (NZF, spanning aa 150-272) and three near the C-terminus (CZF, spanning aa 882-959) of the 1117 amino acid protein (all amino acid numbering corresponds to the mouse sequence). Near the center of the protein is a homeobox domain (aa 559-618) that bears similarity to POU homeodomains in the helical regions, but lacks a basic amino acid cluster usually found on the N-terminal side of the domain. An NMR structure of the isolated Zeb1 homeodomain has been deposited in the Protein Data Bank (PDB 2E19) (Figure 9).
The zinc finger arrays of Zeb1 bind to DNA, but the homeodomain does not (6;7). Each isolated cluster of zinc fingers bound independently to the consensus sequence CAC(C/G)(T/G) (G/T), with highest affinity for CACCT(G) (5-7). These sequences encompass a subset of E boxes (consensus CANNTG), which often occur in tandem and are typically bound by activating transcription factors of the basic helix loop helix (bHLH) family. Some E box-like sequences can negatively regulate gene expression. The isolated Zeb1 NZF and CZF domains appear to have similar binding specificity (5), but the CZF domain had higher binding affinity for target DNA sequences relative to the NZF domain (6). The presence of both intact zinc finger clusters was required for DNA binding and repressor activity in the full Zeb1 protein (6). In another study, Zeb1 was shown to regulate transcription either positively or negatively depending on alternative DNA-binding modes, through either the NZF or CZF domain alone (7). Although the homeodomain of Zeb1 had no specific DNA-binding activity, it has been shown to interact with the NZF domain of Zeb1 itself (7).
Like Zeb1, the 1215 amino acid Zeb2 has an NZF with four individual zinc fingers (three C2H2 and one CCHC), and a CZF with three C2H2 zinc fingers (8). The DNA binding specificity of Zeb2 appears to mirror that of Zeb1 for CACCT sequences. Similarly to Zeb1, the presence of intact NZF and CZF was required for high-affinity binding to DNA and optimal repressor activity (9). Zeb2 has a centrally located homeobox domain that is 42% identical to that of Zeb1.
The existence of a repressor domain in Zeb1 has been substantiated by experiments in which portions of Zeb1 were tethered to heterologous DNA binding domains and tested for their effect on transcription of a reporter construct. However, different groups have attributed repressive activity to distinct regions of the Zeb1 protein. Repressor domains have been identified at the N-terminus (aa 19-127) (6), in a large central region between the two zinc finger clusters (aa 303-902) (10), and in two separate regions within aa 303-902 (aa 302-542 and 760-902) (11). The region between the CZF domain and the C terminus is highly rich in glutamic acid residues (38%), and has been characterized as an activation domain (aa 1011-1124) (12).
Discoveries of interactors have led to the identification of several protein binding domains in Zeb1. (i) Three five-amino acid sequences (PLNLC, PLDLS, PLNLS) between the homeodomain and the CZF domain bind to the co-repressor CtBP, and this region (spanning aa 685-749) is designated the CtBP interaction domain (CID) (13). CtBP exists in a complex containing Zeb1 and Zeb2 and multiple histone-modifying proteins that act to downregulate gene expression (14). (ii) A region between the NZF and homeobox domains of Zeb1 was identified as a Smad interaction domain (aa 377-456) for Smad1, Smad2, and Smad3 (15). Smad transcription factors are activated upon stimulation of receptors for TGF-β or BMP, and translocate to the nucleus to activate transcription of target genes. Zeb1 synergizes with TGF-β/BMP in transcriptional activation by aiding in the recruitment of p300 and P/CAF (histone acetylases) through a direct interaction involving the N-terminal region of Zeb1 (16). (iii) Finally, the negative cofactor NC2 binds to amino acids 726-829 of Zeb1 in yeast two-hybrid assays, and can mediate Zeb1-dependent repression of a reporter in vitro (12). NC2 binds to TATA-binding protein and prevents assembly of the transcription initiation complex (17).
Zeb1 is both phosphorylated and sumoylated. Phosphorylation at serine and threonine residues, but not tyrosine residues, results in two distinctly migrating species upon gel electrophoresis (18). These species are differentially expressed among several cell lines. Zeb1 is sumoylated in vitro on lysine 327 (IK327TE) and lysine 752 (AK752KE) (19), but the effect of this posttranslational modification on Zeb1 function is unknown. Sumoylation of Zeb2 by polycomb protein Pc2 has been shown to attenuate the repressive effect of Zeb2 at the E-cadherin promoter (19).
The cellophane mutation truncates the Zeb1 protein after amino acid 901, resulting in a protein lacking the C-terminal 216 amino acids that encompass part of the CZF domain and the glutamic acid-rich activation domain.
By Northern analysis, Zeb1 mRNA was detected in heart, brain, placenta, and muscle, and at low levels in lung, liver, and kidney (20). In the brain, Zeb1 mRNA was in cerebellum, medulla, spinal cord, occipital lobe, frontal lobe, temporal lobe, and putamen. Among muscle tissues, Zeb1 mRNA was found in skeletal muscle, heart, and in smooth muscle of the uterus, colon, and bladder (20;21).
Immune system tissues expressing Zeb1 mRNA were spleen, lymph node, thymus, and bone marrow (20). However, another group reported Zeb1 transcript in thymocytes and at very low levels in bone marrow cells, but not in splenocytes consisting of only mature B and T cells; they concluded that expression of Zeb1 is downregulated upon migration of cells to the periphery (22). Zeb1 mRNA was detected in the transformed human B cell line Namalwa (5).
By immunohistochemistry and in situ hybridization, the embryonic expression of Zeb1 was observed in mesodermal tissues (e.g., notochord, somite, limb bud mesenchyme, heart); in neural crest derivatives (e.g., dorsal root ganglia, cephalic ganglia); in neuroectoderm; and in parts of the central nervous system (hindbrain, motor neurons in the spinal cord). This pattern was similar in mouse (22;23) and chicken embryos (2), except that Zeb1 was detected in the lens of the eye in chicken but not mouse embryos.
Physiological functions of Zeb1
Zeb1 was discovered independently as δEF1 in chicken (2); Zfh1 in fly (3); Nil-2 (25), ZEB (5), and AREB6 (26) in humans; BZP in hamster (24); δEF1 (27) and MEB1 (4) in mouse; and Zfhep in rat (28). Studies of mutant organisms deficient in Zeb1 demonstrated a role for Zeb1 in embryonic development, and in the differentiation of certain tissues.
In the fly
In the fly, Zeb1 (Zfh-1) is expressed in the mesoderm of early embryos, in a number of mesodermally-derived structures of late embryos, and in many motor neurons of the developing CNS (29). Flies with loss of function mutations of zfh-1 displayed local errors in cell fate or positioning, but normal segregation of the mesoderm and differentiation of mesodermally derived tissues (30). Zfh-1 mutant embryos also showed mild alterations in the number and position of embryonic somatic muscles (30;31). In addition, zfh-1 is required for germ cell migration and gonadal mesoderm development (32), and for formation of a subset of cells in the developing heart (33).
In the mouse
Mutant mice with homozygous null mutation of Zeb1 developed to term but died shortly after birth (23). Overall, Zeb1-/- embryos appeared growth retarded and had short limbs beginning around E15. Shortened distal maxillae and mandibles, curled tails, edema, exencephaly, internal bleeding in the nasal region, and failure of spinal cord closure were observed in embryos at varying frequencies. Many skeletal defects were observed in embryos, including craniofacial abnormalities (e.g. cleft palate, cartilage hyperplasia, dysplasia of nasal septum), limb defects (e.g. shortening and broadening of long bones, fusion of bones and joints), fusion of ribs, sternum defects, and hypoplasia of intervertebral discs. Defects in chondrogenesis may be due in part to dysregulation of the transcription of genes encoding Indian hedgehog (Ihh) (34) and collagen II α1 (Col2a1) (35). Aberrant transcriptional regulation of the collagen I α1 gene in osteoblasts may contribute to the bone abnormalities of Zeb1-/- embryos (36). Zeb1-/- embryos also had small hypocellular thymi with no histological distinction between medulla and cortex, and one tenth the number of thymocytes relative to wild type mice. Female Zeb1+/- mice displayed increased fat mass relative to wild type female mice when fed either regular chow or high-fat chow (37).
In contrast, mice with a truncation of the C terminus of Zeb1 following amino acid 727 (Zeb1ΔC727/ΔC727), and therefore lacking the cluster of C-terminal zinc fingers and the Glu-rich domain, displayed none of the skeletal defects of Zeb1-/- mice (22). Approximately 80% of Zeb1ΔC727/ΔC727 mice died within two days after birth. Like those of Zeb1-/- mice, thymi of surviving Zeb1ΔC727/ΔC727 mice were smaller and contained 0.2% to 1% the number of thymocytes found in wild type mice (22). The medulla and cortex were indistinguishable upon histological analysis of Zeb1ΔC727/ΔC727 thymus sections. Spleen size and cellularity were similar between Zeb1ΔC727/ΔC727 and wild type mice, although there was a trend toward reduced cellularity in Zeb1ΔC727/ΔC727 mice. Lymph node cellularity was 10% of that in wild type mice. Cell populations in thymus, spleen, and lymph node were analyzed by flow cytometry (Table 1). There was a depletion of c-kit+ thymocytes. Of those T cells that reached maturity in Zeb1ΔC727/ΔC727 mice most were CD4+, consistent with the existence of an E-box sequence in a Cd4 enhancer to which Zeb1 binds to repress transcription (38). Forward light scatter analysis showed a significant population of DP thymocytes was larger, and a population of DN thymocytes was smaller in Zeb1ΔC727/ΔC727 mice than in Zeb1ΔC727/+ mice, although the significance of this observation is unknown. Zeb1ΔC727/ΔC727 splenic T cells proliferated normally upon Con A stimulation in vitro.
Table 1. Percentages of cell types in thymus, spleen, and LN of Zeb1ΔC727/ΔC727 mice
Similar numbers of B cells existed in Zeb1ΔC727/ΔC727 and Zeb1ΔC727/+ mice. The population of myeloid cells was reported to be normal in Zeb1ΔC727/ΔC727 mice.
Although no muscle defects have been reported in Zeb1 mutant mice, several groups have provided evidence that Zeb1 regulates muscle cell differentiation. The transcription factor p73 is transcriptionally activated by muscle regulatory factors MyoD, myogenin, Myf5 and Myf6, which bind to E-box sequences of enhancers. Zeb1 competes with these bHLH transcription factors for binding to E-box sites, and has been shown to inhibit MyoD-induced myogenesis in vitro (10;39). Zeb1 also opposes MyoD/Myf5- and MyoD/Myf6-mediated transcriptional activation of p73 by binding to a silencer element in the first intron of p73 (40;41). Zeb1 also negatively regulates integrin α7 expression in myoblasts during skeletal muscle myogenesis by binding to the negative regulatory region in the promoter (42). In vascular smooth muscle cells (SMC), Zeb1 represses expression of collagen I α2 by competing with transcriptional activator Nkx2.5 for binding to an enhancer element (43). Zeb1 also promotes transcription during differentiation of vascular SMC through cooperation with serum response factor (SRF) and Smad3 downstream of TGF-β signaling (21). Zeb1 binds to Smad1, Smad2, and Smad3 (15), and promotes the recruitment of histone acetylases p300 and P/CAF, which act to increase the accessibility of DNA to the transcriptional apparatus (16).
Mechanisms of transcriptional repression and activation
Zeb1 has been shown to repress transcription by several mechanisms (Figure 10). First, it may recruit co-repressors, such as CtBP (14), NC2 (12), and BRG1 (44), to target genes to modify chromatin and histone organization and/or to block assembly of the transcriptional machinery. Second, it competes with transcriptional activators for binding to promoters and/or enhancers, as for example to the Ig heavy chain enhancer (5), the CD4 promoter (38), and the integrin α7 promoter (42). Third, Zeb1 may directly repress transcription through repressor domains located at its N and C termini; mutants lacking one of these repressor domains displayed reduced repressive activity despite retaining DNA binding activity (6;10).
Zeb1 also acts as a transcriptional activator, as has been demonstrated for the genes encoding the vitamin D3 receptor, cyclin G2, and p130 (45;46). Zeb1 can aid in the recruitment of histone acetylases, as mentioned above, and may also directly activate transcription through its activation domain.
Zeb1 in cancer
Through its ability to promote the ‘epithelial to mesenchymal transition’ (EMT), Zeb1 has been implicated in cancer invasion and metastasis. During the EMT, epithelial cells lose polarity and contact with neighboring cells, and adopt a mesenchymal phenotype characterized by the acquisition of migratory and invasive properties (47). This transition is necessary during many developmental processes, such as gastrulation, neural crest formation, and heart development. EMT also plays an important role in tumor progression by supporting tumor cell extension, detachment, and invasion into the adjacent stroma (47). Zeb1 induces EMT in many human cancers, including prostate, colon, breast, and pancreatic cancers, and is known to do so by suppressing the expression of basement membrane components (48) and cell polarity factors (49). Zeb1 also represses the cell adhesion molecule E-cadherin through binding to conserved E-boxes in the E-cadherin promoter [(50), reviewed in (51)]. Downregulation of E-cadherin is a key feature of EMT, and E-cadherin is repressed during malignant transformation. Zeb1 (and Zeb2) are also the most prominent targets of the miR-200 family of microRNAs, which has been shown to revert EMT and induce epithelial differentiation [(52), reviewed in (53)]. miR-200 family members are divided into subgroup I (miR-200a and miR-141) and subgroup II (miR-200b, miR-200c and miR-429). The Zeb1 3’ UTR contains eight miR-200 binding sites, five for subgroup II and three for subgroup I. Conversely, Zeb1 directly inhibits the transcription of miR-200 family microRNAs (54).
Heterozygous null mutations of ZEB1 in humans are linked to posterior polymorphous corneal dystrophy-3 (PPCD3) (OMIM #609141), and have been estimated to cause one-third to one-half of all PPCD cases (55;56). PPCD is an autosomal dominant corneal disorder usually affecting both eyes and characterized by hyperplasia of corneal endothelial cells, which adopt an epithelial phenotype and gene expression pattern, and produce an abnormal basement membrane (Descemet’s membrane). The severity of PPCD symptoms varies, and may include opacities, irodocorneal adhesions (abnormal connection of the iris and cornea), corneal edema, corectopia (displacement of the pupil from its normal, central position) and secondary glaucoma. Mutations in ZEB1 causative for PPCD are associated with abdominal and inguinal hernias (57). In contrast to null mutations, hypomorphic mutations of ZEB1 cause late-onset Fuchs corneal dystrophy, in which guttae form and Descemet’s membrane thickens, leading to corneal edema (58).
Similar to humans with PPCD, Zeb1 homozygous null mouse embryos and heterozygous adult mice displayed thickened corneas with hyperplasia and epithelialization of the endothelium (59). Corneal endothelia of mutant embryos aberrantly expressed epithelial markers including cytokeratin, E-cadherin, and collagen IV α3 (COL4A3) (59). Mutations in human COL4A3, COL4A4, and COL4A5 cause basement membrane abnormalities resulting in the renal disease Alport syndrome (OMIM #301050; #203780) (see aoba, a mouse Col4a4 allele (59)). A binding site for ZEB1 was identified in the promoter of COL4A3, and ectopic expression of COL4A3 was observed in the corneal endothelium of PPCD patients (55). Thus, ZEB1 mutations may result in de-repression of COL4A3 leading to the observed defects in Descemet’s membrane in PPCD.
The cellophane mutation is believed to result in a truncated Zeb1 protein with hypomorphic function. In support of this idea, cellophane mice displayed a phenotype similar to that of Zeb1ΔC727/ΔC727 mice (22) in that their thymi were small and hypocellular, they had fewer DP thymocytes and expanded SP thymocytes, and mature B cell frequencies were normal in spleen and bone marrow. Furthermore, the phenotypes of Zeb1ΔC727/ΔC727 mice and cellophane mice were considerably less severe than that of homozygous null mice, which displayed numerous skeletal defects.
In wild type B cells, Zeb1 mRNA was upregulated within 30 minutes after BCR cross-linking (Figure 11B) (1). This finding suggests that Zeb1 regulates early cellular events following BCR activation. Consistent with this hypothesis, B cells from cellophane mice displayed reduced proliferation in response to BCR crosslinking, although proliferation was normal after treatment with CpG oligonucleotides (Figure 11C and D). We propose that the reduced ability of cellophane B cells to proliferate in response to BCR stimulation leads to impaired antibody responses and GC formation following immunization. More studies are needed to identify the targets of Zeb1 regulation in B cells, and to determine the mechanism by which Zeb1 controls B cell proliferation during humoral immune responses.
|Primers||Primers cannot be located by automatic search.|
Cellophane genotyping is performed by amplifying the region containing the mutation using PCR, followed by sequencing of the amplified region to detect the single nucleotide change.
cellophane (F): 5’-TCAGGTGGAGGGCTTCACATCTAAC -3’
cellophane (R): 5’- GCTCTGTCAGCATAGACACCAAGG -3’
1) 95°C 2:00
2) 95°C 0:30
3) 56°C 0:30
4) 72°C 1:00
5) repeat steps (2-4) 29X
6) 72°C 7:00
7) 4°C ∞
Primers for sequencing
cellophane_seq(F): 5’- GTGTAGATCCCAGGGATTCAACTC -3’
cellophane_seq(R): 5’- CACCAAGGCATTAAAGGCG -3’
The following sequence of 670 nucleotides (from Genbank genomic region NC_000084.5 for linear genomic sequence of Zeb1, sense strand) is amplified:
178206 tcagg tggagggctt cacatctaac atgaaatttc tctcttgtgc atctcttgac
178261 tttcctacat ttattgactt attggcagtg ggtgctttgt gtgccagctc tcacacgtgg
178321 aggtccgagg acaacttgtg ggagtcagtt ctctcctttc accgtgtaga tcccagggat
178381 tcaactctgc tcatctttct ttgtggcaaa tagctttgtc tgctgagcca tctcagtgac
178441 cttatctctt gggaatcttt ttctttgaca tttaatcttc tttttccact taggatgaaa
178501 gacaagacac tagctcagaa ggagtctcca ctgtggagga ccagaatgac tctgactcca
178561 cgccacccaa aaagaaaact cggaagacag agaatggaat gtatgcatgt gacctgtgtg
178621 acaagatatt tcagaagagc agctcactgt tgagacacaa atatgagcac acaggtgtgt
178681 gggggacctg ggcacgaggt tctaaaggtg cctgtggccc agtgcacatg aaacatgccc
178741 atagtgtgtg acgttctcag ctctgctccg tggtctccac ttttcacata ttcctactgc
178801 ggagcagtgg cttggctgtc tgctgcctag actcccgttt cgcctttaat gccttggtgt
178861 ctatgctgac agagc
Primer binding sites are underlined; sequencing primer binding sites are highlighted in gray; the mutated T is indicated in red.
1. Arnold, C. N., Pirie, E., Dosenovic, P., McInerney, G. M., Xia, Y., Wang, N., Li, X., Siggs, O. M., Karlsson Hedestam, G. B., and Beutler, B. (2012) A Forward Genetic Screen Reveals Roles for Nfkbid, Zeb1, and Ruvbl2 in Humoral Immunity. Proc. Natl. Acad. Sci. U. S. A.. .
2. Funahashi, J., Sekido, R., Murai, K., Kamachi, Y., and Kondoh, H. (1993) Delta-Crystallin Enhancer Binding Protein Delta EF1 is a Zinc Finger-Homeodomain Protein Implicated in Postgastrulation Embryogenesis. Development. 119, 433-446.
3. Fortini, M. E., Lai, Z. C., and Rubin, G. M. (1991) The Drosophila Zfh-1 and Zfh-2 Genes Encode Novel Proteins Containing both Zinc-Finger and Homeodomain Motifs. Mech. Dev.. 34, 113-122.
4. Genetta, T., and Kadesch, T. (1996) Cloning of a cDNA Encoding a Mouse Transcriptional Repressor Displaying Striking Sequence Conservation Across Vertebrates. Gene. 169, 289-290.
5. Genetta, T., Ruezinsky, D., and Kadesch, T. (1994) Displacement of an E-Box-Binding Repressor by Basic Helix-Loop-Helix Proteins: Implications for B-Cell Specificity of the Immunoglobulin Heavy-Chain Enhancer. Mol. Cell. Biol.. 14, 6153-6163.
6. Sekido, R., Murai, K., Kamachi, Y., and Kondoh, H. (1997) Two Mechanisms in the Action of Repressor deltaEF1: Binding Site Competition with an Activator and Active Repression. Genes Cells. 2, 771-783.
7. Ikeda, K., and Kawakami, K. (1995) DNA Binding through Distinct Domains of Zinc-Finger-Homeodomain Protein AREB6 has Different Effects on Gene Transcription. Eur. J. Biochem.. 233, 73-82.
8. Verschueren, K., Remacle, J. E., Collart, C., Kraft, H., Baker, B. S., Tylzanowski, P., Nelles, L., Wuytens, G., Su, M. T., Bodmer, R., Smith, J. C., and Huylebroeck, D. (1999) SIP1, a Novel Zinc finger/homeodomain Repressor, Interacts with Smad Proteins and Binds to 5'-CACCT Sequences in Candidate Target Genes. J. Biol. Chem.. 274, 20489-20498.
9. Remacle, J. E., Kraft, H., Lerchner, W., Wuytens, G., Collart, C., Verschueren, K., Smith, J. C., and Huylebroeck, D. (1999) New Mode of DNA Binding of Multi-Zinc Finger Transcription Factors: DeltaEF1 Family Members Bind with Two Hands to Two Target Sites. EMBO J.. 18, 5073-5084.
10. Postigo, A. A., and Dean, D. C. (1997) ZEB, a Vertebrate Homolog of Drosophila Zfh-1, is a Negative Regulator of Muscle Differentiation. EMBO J.. 16, 3935-3943.
11. Postigo, A. A., and Dean, D. C. (1999) Independent Repressor Domains in ZEB Regulate Muscle and T-Cell Differentiation. Mol. Cell. Biol.. 19, 7961-7971.
12. Ikeda, K., Halle, J. P., Stelzer, G., Meisterernst, M., and Kawakami, K. (1998) Involvement of Negative Cofactor NC2 in Active Repression by Zinc Finger-Homeodomain Transcription Factor AREB6. Mol. Cell. Biol.. 18, 10-18.
13. Postigo, A. A., and Dean, D. C. (1999) ZEB Represses Transcription through Interaction with the Corepressor CtBP. Proc. Natl. Acad. Sci. U. S. A.. 96, 6683-6688.
14. Shi, Y., Sawada, J., Sui, G., Affar el, B., Whetstine, J. R., Lan, F., Ogawa, H., Luke, M. P., Nakatani, Y., and Shi, Y. (2003) Coordinated Histone Modifications Mediated by a CtBP Co-Repressor Complex. Nature. 422, 735-738.
15. Postigo, A. A. (2003) Opposing Functions of ZEB Proteins in the Regulation of the TGFbeta/BMP Signaling Pathway. EMBO J.. 22, 2443-2452.
16. Postigo, A. A., Depp, J. L., Taylor, J. J., and Kroll, K. L. (2003) Regulation of Smad Signaling through a Differential Recruitment of Coactivators and Corepressors by ZEB Proteins. EMBO J.. 22, 2453-2462.
17. Kim, T. K., Zhao, Y., Ge, H., Bernstein, R., and Roeder, R. G. (1995) TATA-Binding Protein Residues Implicated in a Functional Interplay between Negative Cofactor NC2 (Dr1) and General Factors TFIIA and TFIIB. J. Biol. Chem.. 270, 10976-10981.
18. Costantino, M. E., Stearman, R. P., Smith, G. E., and Darling, D. S. (2002) Cell-Specific Phosphorylation of Zfhep Transcription Factor. Biochem. Biophys. Res. Commun.. 296, 368-373.
19. Long, J., Zuo, D., and Park, M. (2005) Pc2-Mediated Sumoylation of Smad-Interacting Protein 1 Attenuates Transcriptional Repression of E-Cadherin. J. Biol. Chem.. 280, 35477-35489.
20. Postigo, A. A., and Dean, D. C. (2000) Differential Expression and Function of Members of the Zfh-1 Family of Zinc finger/homeodomain Repressors. Proc. Natl. Acad. Sci. U. S. A.. 97, 6391-6396.
21. Nishimura, G., Manabe, I., Tsushima, K., Fujiu, K., Oishi, Y., Imai, Y., Maemura, K., Miyagishi, M., Higashi, Y., Kondoh, H., and Nagai, R. (2006) DeltaEF1 Mediates TGF-Beta Signaling in Vascular Smooth Muscle Cell Differentiation. Dev. Cell.. 11, 93-104.
22. Higashi, Y., Moribe, H., Takagi, T., Sekido, R., Kawakami, K., Kikutani, H., and Kondoh, H. (1997) Impairment of T Cell Development in deltaEF1 Mutant Mice. J. Exp. Med.. 185, 1467-1479.
23. Takagi, T., Moribe, H., Kondoh, H., and Higashi, Y. (1998) DeltaEF1, a Zinc Finger and Homeodomain Transcription Factor, is Required for Skeleton Patterning in Multiple Lineages. Development. 125, 21-31.
24. Franklin, A. J., Jetton, T. L., Shelton, K. D., and Magnuson, M. A. (1994) BZP, a Novel Serum-Responsive Zinc Finger Protein that Inhibits Gene Transcription. Mol. Cell. Biol.. 14, 6773-6788.
25. Williams, T. M., Moolten, D., Burlein, J., Romano, J., Bhaerman, R., Godillot, A., Mellon, M., Rauscher, F. J.,3rd, and Kant, J. A. (1991) Identification of a Zinc Finger Protein that Inhibits IL-2 Gene Expression. Science. 254, 1791-1794.
26. Watanabe, Y., Kawakami, K., Hirayama, Y., and Nagano, K. (1993) Transcription Factors Positively and Negatively Regulating the Na,K-ATPase Alpha 1 Subunit Gene. J. Biochem.. 114, 849-855.
27. Sekido, R., Takagi, T., Okanami, M., Moribe, H., Yamamura, M., Higashi, Y., and Kondoh, H. (1996) Organization of the Gene Encoding Transcriptional Repressor deltaEF1 and Cross-Species Conservation of its Domains. Gene. 173, 227-232.
28. Cabanillas, A. M., and Darling, D. S. (1996) Alternative Splicing Gives Rise to Two Isoforms of Zfhep, a Zinc finger/homeodomain Protein that Binds T3-Response Elements. DNA Cell Biol.. 15, 643-651.
29. Lai, Z. C., Fortini, M. E., and Rubin, G. M. (1991) The Embryonic Expression Patterns of Zfh-1 and Zfh-2, Two Drosophila Genes Encoding Novel Zinc-Finger Homeodomain Proteins. Mech. Dev.. 34, 123-134.
30. Lai, Z. C., Rushton, E., Bate, M., and Rubin, G. M. (1993) Loss of Function of the Drosophila Zfh-1 Gene Results in Abnormal Development of Mesodermally Derived Tissues. Proc. Natl. Acad. Sci. U. S. A.. 90, 4122-4126.
31. Postigo, A. A., Ward, E., Skeath, J. B., and Dean, D. C. (1999) Zfh-1, the Drosophila Homologue of ZEB, is a Transcriptional Repressor that Regulates Somatic Myogenesis. Mol. Cell. Biol.. 19, 7255-7263.
32. Broihier, H. T., Moore, L. A., Van Doren, M., Newman, S., and Lehmann, R. (1998) Zfh-1 is Required for Germ Cell Migration and Gonadal Mesoderm Development in Drosophila. Development. 125, 655-666.
33. Su, M. T., Fujioka, M., Goto, T., and Bodmer, R. (1999) The Drosophila Homeobox Genes Zfh-1 and Even-Skipped are Required for Cardiac-Specific Differentiation of a Numb-Dependent Lineage Decision. Development. 126, 3241-3251.
34. Bellon, E., Luyten, F. P., and Tylzanowski, P. (2009) Delta-EF1 is a Negative Regulator of Ihh in the Developing Growth Plate. J. Cell Biol.. 187, 685-699.
35. Murray, D., Precht, P., Balakir, R., and Horton, W. E.,Jr. (2000) The Transcription Factor deltaEF1 is Inversely Expressed with Type II Collagen mRNA and can Repress Col2a1 Promoter Activity in Transfected Chondrocytes. J. Biol. Chem.. 275, 3610-3618.
36. Terraz, C., Toman, D., Delauche, M., Ronco, P., and Rossert, J. (2001) Delta Ef1 Binds to a Far Upstream Sequence of the Mouse Pro-Alpha 1(I) Collagen Gene and Represses its Expression in Osteoblasts. J. Biol. Chem.. 276, 37011-37019.
37. Saykally, J. N., Dogan, S., Cleary, M. P., and Sanders, M. M. (2009) The ZEB1 Transcription Factor is a Novel Repressor of Adiposity in Female Mice. PLoS One. 4, e8460.
38. Brabletz, T., Jung, A., Hlubek, F., Lohberg, C., Meiler, J., Suchy, U., and Kirchner, T. (1999) Negative Regulation of CD4 Expression in T Cells by the Transcriptional Repressor ZEB. Int. Immunol.. 11, 1701-1708.
39. Sekido, R., Murai, K., Funahashi, J., Kamachi, Y., Fujisawa-Sehara, A., Nabeshima, Y., and Kondoh, H. (1994) The Delta-Crystallin Enhancer-Binding Protein Delta EF1 is a Repressor of E2-Box-Mediated Gene Activation. Mol. Cell. Biol.. 14, 5692-5700.
40. Fontemaggi, G., Gurtner, A., Damalas, A., Costanzo, A., Higashi, Y., Sacchi, A., Strano, S., Piaggio, G., and Blandino, G. (2005) DeltaEF1 Repressor Controls Selectively p53 Family Members during Differentiation. Oncogene. 24, 7273-7280.
41. Fontemaggi, G., Gurtner, A., Strano, S., Higashi, Y., Sacchi, A., Piaggio, G., and Blandino, G. (2001) The Transcriptional Repressor ZEB Regulates p73 Expression at the Crossroad between Proliferation and Differentiation. Mol. Cell. Biol.. 21, 8461-8470.
42. Jethanandani, P., and Kramer, R. H. (2005) Alpha7 Integrin Expression is Negatively Regulated by deltaEF1 during Skeletal Myogenesis. J. Biol. Chem.. 280, 36037-36046.
43. Ponticos, M., Partridge, T., Black, C. M., Abraham, D. J., and Bou-Gharios, G. (2004) Regulation of Collagen Type I in Vascular Smooth Muscle Cells by Competition between Nkx2.5 and deltaEF1/ZEB1. Mol. Cell. Biol.. 24, 6151-6161.
44. Sanchez-Tillo, E., Lazaro, A., Torrent, R., Cuatrecasas, M., Vaquero, E. C., Castells, A., Engel, P., and Postigo, A. (2010) ZEB1 Represses E-Cadherin and Induces an EMT by Recruiting the SWI/SNF Chromatin-Remodeling Protein BRG1. Oncogene. 29, 3490-3500.
45. Chen, J., Yusuf, I., Andersen, H. M., and Fruman, D. A. (2006) FOXO Transcription Factors Cooperate with Delta EF1 to Activate Growth Suppressive Genes in B Lymphocytes. J. Immunol.. 176, 2711-2721.
46. Lazarova, D. L., Bordonaro, M., and Sartorelli, A. C. (2001) Transcriptional Regulation of the Vitamin D(3) Receptor Gene by ZEB. Cell Growth Differ.. 12, 319-326.
47. Thiery, J. P., Acloque, H., Huang, R. Y., and Nieto, M. A. (2009) Epithelial-Mesenchymal Transitions in Development and Disease. Cell. 139, 871-890.
48. Spaderna, S., Schmalhofer, O., Wahlbuhl, M., Dimmler, A., Bauer, K., Sultan, A., Hlubek, F., Jung, A., Strand, D., Eger, A., Kirchner, T., Behrens, J., and Brabletz, T. (2008) The Transcriptional Repressor ZEB1 Promotes Metastasis and Loss of Cell Polarity in Cancer. Cancer Res.. 68, 537-544.
49. Aigner, K., Dampier, B., Descovich, L., Mikula, M., Sultan, A., Schreiber, M., Mikulits, W., Brabletz, T., Strand, D., Obrist, P., Sommergruber, W., Schweifer, N., Wernitznig, A., Beug, H., Foisner, R., and Eger, A. (2007) The Transcription Factor ZEB1 (deltaEF1) Promotes Tumour Cell Dedifferentiation by Repressing Master Regulators of Epithelial Polarity. Oncogene. 26, 6979-6988.
50. Grooteclaes, M. L., and Frisch, S. M. (2000) Evidence for a Function of CtBP in Epithelial Gene Regulation and Anoikis. Oncogene. 19, 3823-3828.
51. Vandewalle, C., Van Roy, F., and Berx, G. (2009) The Role of the ZEB Family of Transcription Factors in Development and Disease. Cell Mol. Life Sci.. 66, 773-787.
52. Wellner, U., Schubert, J., Burk, U. C., Schmalhofer, O., Zhu, F., Sonntag, A., Waldvogel, B., Vannier, C., Darling, D., zur Hausen, A., Brunton, V. G., Morton, J., Sansom, O., Schuler, J., Stemmler, M. P., Herzberger, C., Hopt, U., Keck, T., Brabletz, S., and Brabletz, T. (2009) The EMT-Activator ZEB1 Promotes Tumorigenicity by Repressing Stemness-Inhibiting microRNAs. Nat. Cell Biol.. 11, 1487-1495.
53. Brabletz, S., and Brabletz, T. (2010) The ZEB/miR-200 Feedback Loop--a Motor of Cellular Plasticity in Development and Cancer? EMBO Rep.. 11, 670-677.
54. Bracken, C. P., Gregory, P. A., Kolesnikoff, N., Bert, A. G., Wang, J., Shannon, M. F., and Goodall, G. J. (2008) A Double-Negative Feedback Loop between ZEB1-SIP1 and the microRNA-200 Family Regulates Epithelial-Mesenchymal Transition. Cancer Res.. 68, 7846-7854.
55. Krafchak, C. M., Pawar, H., Moroi, S. E., Sugar, A., Lichter, P. R., Mackey, D. A., Mian, S., Nairus, T., Elner, V., Schteingart, M. T., Downs, C. A., Kijek, T. G., Johnson, J. M., Trager, E. H., Rozsa, F. W., Mandal, M. N., Epstein, M. P., Vollrath, D., Ayyagari, R., Boehnke, M., and Richards, J. E. (2005) Mutations in TCF8 Cause Posterior Polymorphous Corneal Dystrophy and Ectopic Expression of COL4A3 by Corneal Endothelial Cells. Am. J. Hum. Genet.. 77, 694-708.
56. Liskova, P., Tuft, S. J., Gwilliam, R., Ebenezer, N. D., Jirsova, K., Prescott, Q., Martincova, R., Pretorius, M., Sinclair, N., Boase, D. L., Jeffrey, M. J., Deloukas, P., Hardcastle, A. J., Filipec, M., and Bhattacharya, S. S. (2007) Novel Mutations in the ZEB1 Gene Identified in Czech and British Patients with Posterior Polymorphous Corneal Dystrophy. Hum. Mutat.. 28, 638.
57. Aldave, A. J., Yellore, V. S., Yu, F., Bourla, N., Sonmez, B., Salem, A. K., Rayner, S. A., Sampat, K. M., Krafchak, C. M., and Richards, J. E. (2007) Posterior Polymorphous Corneal Dystrophy is Associated with TCF8 Gene Mutations and Abdominal Hernia. Am. J. Med. Genet. A.. 143A, 2549-2556.
58. Riazuddin, S. A., Zaghloul, N. A., Al-Saif, A., Davey, L., Diplas, B. H., Meadows, D. N., Eghrari, A. O., Minear, M. A., Li, Y. J., Klintworth, G. K., Afshari, N., Gregory, S. G., Gottsch, J. D., and Katsanis, N. (2010) Missense Mutations in TCF8 Cause Late-Onset Fuchs Corneal Dystrophy and Interact with FCD4 on Chromosome 9p. Am. J. Hum. Genet.. 86, 45-53.
|Science Writers||Eva Marie Y. Moresco, Anne Murray|
|Illustrators||Diantha La Vine|
|Authors||Carrie N. Arnold, Elaine Pirie, Bruce Beutler|