Schwann cell development and peripheral nerve myelination require the serial expression of transcriptional activators, such as Sox10, Oct6 (also called Scip or Pou3f1) and Krox20 (also called Egr2). Here we show that transcriptional repression, mediated by the zinc-finger protein Zeb2 (also known as Sip1), is essential for differentiation and myelination. Mice lacking Zeb2 in Schwann cells develop a severe peripheral neuropathy, caused by failure of axonal sorting and virtual absence of myelin membranes. Zeb2-deficient Schwann cells continuously express repressors of lineage progression. Moreover, genes for negative regulators of maturation such as Sox2 and Ednrb emerge as Zeb2 target genes, supporting its function as an 'inhibitor of inhibitors' in myelination control. When Zeb2 is deleted in adult mice, Schwann cells readily dedifferentiate following peripheral nerve injury and become repair cells. However, nerve regeneration and remyelination are both perturbed, demonstrating that Zeb2, although undetectable in adult Schwann cells, has a latent function throughout life.
This is a preview of subscription content, access via your institution
Open Access articles citing this article.
Lessons from Injury: How Nerve Injury Studies Reveal Basic Biological Mechanisms and Therapeutic Opportunities for Peripheral Nerve Diseases
Neurotherapeutics Open Access 30 September 2021
Cancer and Metastasis Reviews Open Access 19 April 2021
Schwann cell plasticity regulates neuroblastic tumor cell differentiation via epidermal growth factor-like protein 8
Nature Communications Open Access 12 March 2021
Subscribe to Journal
Get full journal access for 1 year
only $6.58 per issue
All prices are NET prices.
VAT will be added later in the checkout.
Tax calculation will be finalised during checkout.
Get time limited or full article access on ReadCube.
All prices are NET prices.
Gene Expression Omnibus
Jessen, K.R. & Mirsky, R. Negative regulation of myelination: relevance for development, injury, and demyelinating disease. Glia 56, 1552–1565 (2008).
Monk, K.R., Feltri, M.L. & Taveggia, C. New insights on Schwann cell development. Glia 63, 1376–1393 (2015).
Stolt, C.C. & Wegner, M. Schwann cells and their transcriptional network: evolution of key regulators of peripheral myelination. Brain Res. http://dx.doi.org/10.1016/j.brainres.2015.09.025 (2015).
Topilko, P. et al. Krox-20 controls myelination in the peripheral nervous system. Nature 371, 796–799 (1994).
Decker, L. et al. Peripheral myelin maintenance is a dynamic process requiring constant Krox20 expression. J. Neurosci. 26, 9771–9779 (2006).
Ghislain, J. & Charnay, P. Control of myelination in Schwann cells: a Krox20 cis-regulatory element integrates Oct6, Brn2 and Sox10 activities. EMBO Rep. 7, 52–58 (2006).
Kuhlbrodt, K., Herbarth, B., Sock, E., Hermans-Borgmeyer, I. & Wegner, M. Sox10, a novel transcriptional modulator in glial cells. J. Neurosci. 18, 237–250 (1998).
Britsch, S. et al. The transcription factor Sox10 is a key regulator of peripheral glial development. Genes Dev. 15, 66–78 (2001).
Finzsch, M. et al. Sox10 is required for Schwann cell identity and progression beyond the immature Schwann cell stage. J. Cell Biol. 189, 701–712 (2010).
Fröb, F. et al. Establishment of myelinating Schwann cells and barrier integrity between central and peripheral nervous systems depend on Sox10. Glia 60, 806–819 (2012).
Bremer, M. et al. Sox10 is required for Schwann-cell homeostasis and myelin maintenance in the adult peripheral nerve. Glia 59, 1022–1032 (2011).
Le, N. et al. Nab proteins are essential for peripheral nervous system myelination. Nat. Neurosci. 8, 932–940 (2005).
Desmazières, A., Decker, L., Vallat, J.M., Charnay, P. & Gilardi-Hebenstreit, P. Disruption of Krox20-Nab interaction in the mouse leads to peripheral neuropathy with biphasic evolution. J. Neurosci. 28, 5891–5900 (2008).
Mager, G.M. et al. Active gene repression by the Egr2.NAB complex during peripheral nerve myelination. J. Biol. Chem. 283, 18187–18197 (2008).
He, Y. et al. The transcription factor Yin Yang 1 is essential for oligodendrocyte progenitor differentiation. Neuron 55, 217–230 (2007).
He, Y. et al. Yy1 as a molecular link between neuregulin and transcriptional modulation of peripheral myelination. Nat. Neurosci. 13, 1472–1480 (2010).
Verschueren, K. et al. 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 (1999).
Conidi, A. et al. Few Smad proteins and many Smad-interacting proteins yield multiple functions and action modes in TGFβ/BMP signaling in vivo. Cytokine Growth Factor Rev. 22, 287–300 (2011).
Comijn, J. et al. The two-handed E box binding zinc finger protein SIP1 downregulates E-cadherin and induces invasion. Mol. Cell 7, 1267–1278 (2001).
Vandewalle, C. et al. SIP1/ZEB2 induces EMT by repressing genes of different epithelial cell-cell junctions. Nucleic Acids Res. 33, 6566–6578 (2005).
Dai, Y.-H. et al. ZEB2 promotes the metastasis of gastric cancer and modulates epithelial mesenchymal transition of gastric cancer cells. Dig. Dis. Sci. 57, 1253–1260 (2012).
Seuntjens, E. et al. Sip1 regulates sequential fate decisions by feedback signaling from postmitotic neurons to progenitors. Nat. Neurosci. 12, 1373–1380 (2009).
Weng, Q. et al. Dual-mode modulation of Smad signaling by Smad-interacting protein Sip1 is required for myelination in the central nervous system. Neuron 73, 713–728 (2012).
Van de Putte, T., Francis, A., Nelles, L., van Grunsven, L.A. & Huylebroeck, D. Neural crest-specific removal of Zfhx1b in mouse leads to a wide range of neurocristopathies reminiscent of Mowat-Wilson syndrome. Hum. Mol. Genet. 16, 1423–1436 (2007).
Mowat, D.R. et al. Hirschsprung disease, microcephaly, mental retardation, and characteristic facial features: delineation of a new syndrome and identification of a locus at chromosome 2q22-q23. J. Med. Genet. 35, 617–623 (1998).
Stanchina, L., Van de Putte, T., Goossens, M., Huylebroeck, D. & Bondurand, N. Genetic interaction between Sox10 an Zfhx1b during enteric nervous sstem development. Dev. Biol. 341, 416–428 (2010).
Higashi, Y. et al. Generation of the floxed allele of the SIP1 (Smad-interacting protein 1) gene for Cre-mediated conditional knockout in the mouse. Genesis 32, 82–84 (2002).
Jaegle, M. et al. The POU proteins Brn-2 and Oct-6 share important functions in Schwann cell development. Genes Dev. 17, 1380–1391 (2003).
Jessen, K.R. & Mirsky, R. The origin and development of glial cells in peripheral nerves. Nat. Rev. Neurosci. 6, 671–682 (2005).
Colognato, H. & Tzvetanova, I.D. Glia unglued: how signals from the extracellular matrix regulate the development of myelinating glia. Dev. Neurobiol. 71, 924–955 (2011).
Brennan, A. et al. Endothelins control the timing of Schwann cell generation in vitro and in vivo. Dev. Biol. 227, 545–557 (2000).
Druckenbrod, N.R., Powers, P.A., Bartley, C.R., Walker, J.W. & Epstein, M. L. Targeting of endothelin receptor-B to the neural crest. Genesis 46, 396–400 (2008).
Xin, M. et al. Essential roles of the bHLH transcription factor Hrt2 in repression of atrial gene expression and maintenance of postnatal cardiac function. Proc. Natl. Acad. Sci. USA 104, 7975–7980 (2007).
Leone, D.P. et al. Tamoxifen-inducible glia-specific Cre mice for somatic mutagenesis in oligodendrocytes and Schwann cells. Mol. Cell. Neurosci. 22, 430–440 (2003).
Madisen, L. et al. A robust and high-throughput Cre reporting and characterization system for the whole mouse brain. Nat. Neurosci. 13, 133–140 (2010).
Wu, L.M.N. et al. Zeb2 recruits HDAC-NuRD to inhibit Notch and controls Schwann cell differentiation and remyelination. Nat. Neurosci. http://dx.doi.org/10.1038/nn.4322 (2016).
Weider, M. et al. Chromatin-remodeling factor Brg1 is required for Schwann cell differentiation and myelination. Dev. Cell 23, 193–201 (2012).
Bermingham, J.R. Jr. et al. Tst-1/Oct-6/SCIP regulates a unique step in peripheral myelination and is required for normal respiration. Genes Dev. 10, 1751–1762 (1996).
Jaegle, M. et al. The POU factor Oct-6 and Schwann cell differentiation. Science 273, 507–510 (1996).
Verstappen, G. et al. Atypical Mowat-Wilson patient confirms the importance of the novel association between ZFHX1B/SIP1 and NuRD corepressor complex. Hum. Mol. Genet. 17, 1175–1183 (2008).
Berti, C. et al. Non-redundant function of dystroglycan and β1 integrins in radial sorting of axons. Development 138, 4025–4037 (2011).
Yang, D. et al. Coordinate control of axon defasciculation and myelination by laminin-2 and -8. J. Cell Biol. 168, 655–666 (2005).
Le, N. et al. Analysis of congenital hypomyelinating Egr2Lo/Lo nerves identifies Sox2 as an inhibitor of Schwann cell differentiation and myelination. Proc. Natl. Acad. Sci. USA 102, 2596–2601 (2005).
Weber, D. et al. Mechanisms of epigenetic and cell-type specific regulation of Hey target genes in ES cells and cardiomyocytes. J. Mol. Cell. Cardiol. 79, 79–88 (2015).
Woodhoo, A. et al. Notch controls embryonic Schwann cell differentiation, postnatal myelination and adult plasticity. Nat. Neurosci. 12, 839–847 (2009).
Evans, E. et al. The behavioral phenotype of Mowat-Wilson syndrome. Am. J. Med. Genet. A 158A, 358–366 (2012).10.1002/ajmg.a.34405
Pradier, B. et al. Smad-interacting protein 1 affects acute and tonic, but not chronic pain. Eur. J. Pain 18, 249–257 (2014).
Emery, B. Regulation of oligodendrocyte differentiation and myelination. Science 330, 779–782 (2010).
Li, H., He, Y., Richardson, W.D. & Casaccia, P. Two-tier transcriptional control of oligodendrocyte differentiation. Curr. Opin. Neurobiol. 19, 479–485 (2009).
Arthur-Farraj, P.J. et al. c-Jun reprograms Schwann cells of injured nerves to generate a repair cell essential for regeneration. Neuron 75, 633–647 (2012).
Adameyko, I. et al. Schwann cell precursors from nerve innervation are a cellular origin of melanocytes in skin. Cell 139, 366–379 (2009).
Toda, K., Small, J.A., Goda, S. & Quarles, R.H. Biochemical and cellular properties of three immortalized Schwann cell lines expressing different levels of the myelin-associated glycoprotein. J. Neurochem. 63, 1646–1657 (1994).
Klapdor, K., Dulfer, B.G., Hammann, A. & Van der Staay, F.J. A low-cost method to analyse footprint patterns. J. Neurosci. Methods 75, 49–54 (1997).
Inserra, M.M., Bloch, D.A. & Terris, D.J. Functional indices for sciatic, peroneal, and posterior tibial nerve lesions in the mouse. Microsurgery 18, 119–124 (1998).
Darbas, A. et al. Cell autonomy of the mouse claw paw mutation. Dev. Biol. 272, 470–482 (2004).
The authors would like to thank C. Maack, T. Durkaya and A. Fahrenholz for excellent technical assistance. We thank the staff of the Transcriptome Analysis Laboratory (TAL) of the University Medical Center Göttingen for performance and statistical analysis of the microarray analysis and A. Diedrich, M. Wehe, B. Nickel and T. Hoffmeister for excellent support in animal husbandry. We are grateful to Q.R. Lu for communicating unpublished data. M.W.S. is supported by a Heisenberg Professorship granted by the DFG (GZ: SE 1944/1-1). D.H. is supported by Belspo-IAP funding (IAPVII-07), FWO-V (G.0782.14), Hercules Foundation (ZW09-03 project InfraMouse) and Erasmus MC start-up funds. K.-A.N. is supported by the DFG (Research Center Molecular Physiology of the Brain, CNMPB) and holds a European Research Council Advanced Investigator Grant.
The authors declare no competing financial interests.
Integrated supplementary information
(a) The number of Schwann cell nuclei per cross section of sciatic nerve was not different between conditional single mutants (Dhh-cre::Zeb2fl/fl) and both conditional double mutants (Dhh-cre::Zeb2fl/fl::Ednrbfl/fl and Dhh-cre::Zeb2fl/fl::Hey2fl/fl) at age P25. At age P365, the Schwann cell number of Dhh-cre::Zeb2fl/fl mice was not significantly different from P25. Bars, mean ±SD (n=3 animals per age and genotype, P=0.0522, H=7.717949, Kruskal-Wallis one-way ANOVA).
(b) Unaltered cell proliferation in sciatic nerve as determined by incorporation of bromo-desoxyuridine (BrdU) in Dhh-cre::Zeb2fl/fl mice compared to controls at E18.5, P10 and P25. Values are expressed as the percentage (±SD) of BrdU-positive cells of all DAPI-positive nuclei. (n=4 mice per genotype, except for the P10 time point n=3, two-sided Student’s t-test of unpaired samples E18.5 P=0.339, t=1.039350, P10 P=0.875, t=0.1679966, P25 P=0.697, t=0.4061983).
(c) Semi-thin sections of controls and mutants at age 1 year showing comparable size of sciatic nerve branches. Scale bar 50 µm.
(d) The number of axons counted in 15 randomly taken electron micrographs is not significantly different in mutants and controls. Bars, mean ±SD (n=3 animals per genotype, age 1 year, two-sided Student’s t-test of unpaired samples, P=0.192, t=1.568639).
(a) In vivo chromatin immunoprecipitation of Zeb2 targets using DNA from pooled sciatic nerves of mice at age P1 (n=3 individual preparations of genomic DNA from 15 mice at age P1 in each preparation). Fold enrichment (by qPCR) of the indicated promoter fragments relative to control samples (no antibody) is shown for three independent experiments (variance introduced at the step of shearing pooled genomic DNA). A region in the E-cadherin promoter (Cdh, a known target gene of Zeb2) without Zeb2 binding site served as an independent negative control (neg). The location of the different primer pairs (#1 to #9) is shown schematically in (b).
Analyzed 4 weeks after the last injection of tamoxifen, induced tdTomato fluorescence (red) was observed in cryostat cross sections of sciatic nerves from PLPCreERT2::Zeb2fl/fl mice (top panel), but not in tamoxifen-treated Zeb2fl/fl control mice (bottom panel). Green: myelin (fluoromyelin). Red: tdTomato. Scale bar, 10 µm (Representative images of n=3 mice per genoytpe).
(a) Myelin sheath thickness was unaltered 12 weeks after the last injection of tamoxifen in PLP-creERT2::Zeb2fl/fl mice, when compared to the 3 respective control groups. Representative images of 3 animals per group. Scale bar, 2.5 µm.
(b) Scatter plot and g-ratio analysis of tamoxifen-treated PLP-creERT2::Zeb2fl/fl mice in comparison to tamoxifen-treated Zeb2fl/fl mice (n=3 animals per group, at least 100 randomly chosen fibers per animal, two-sided Student’s t-test of unpaired samples, P=0.504, t=0.7339136).
(a) Semi-thin sections of sciatic nerves from tamoxifen-treated Zeb2fl/fl and PLP-creERT2::Zeb2fl/fl mice 11 and 28 days after nerve crush. Note the reduced number of remyelinated fibers in nerves of PLP-creERT2::Zeb2fl/fl mice 28 days after injury (scale bar 20 µm).
(b) Scatter plot and G-ratio analysis of remyelinated fibers 8 weeks after nerve crush. Note that once remyelination occured resulting myelin sheath thickness was not significantly different in tamoxifen-treated PLP-creERT2::Zeb2fl/fl mice and the 3 respective control groups (n=3 animals per group, at least 100 randomly chosen fibers per animal, Kruskal-Wallis one-way ANOVA P=0.875, H=0.6923077).
Schwann cell (SC) differentiation is simplified in a linear order from SC precursors (left) to immature SC, to promyelin stage SC, and myelinating SC (right). Various promyelinating transcription factors drive differentiation (top: boxed in light green), whereas developmental inhibitors (bottom: boxed in light red) antagonize this process. In both groups, transcriptional activators (green fonts) and transcriptional repressors (red fonts) are found. Zeb2 is expressed throughout the early Schwann cell lineage and is a repressor of genes encoding developmental inhibitors ("inhibiting the inhibitors"), thereby promoting differentiation. Zeb2 inactivation in the Schwann cell lineage leads to continous expression of developmental inhibitors, such as Sox2, Hey2 and Ednrb (selected for analysis in this study), causing a developmental arrest of Zeb2-deficient SC prior to axonal sorting. When healthy adult nerves are injured, c-Jun drives SC dedifferentiation and the formation of 'repair cells', which is also possible in Zeb2 mutant mice. However, efficient redifferentiation to myelination-competent Schwann cells is Zeb2-dependent and often times fails in adult conditional Zeb2 mutants. For clarity, only a subset of known factors is depicted.
About this article
Cite this article
Quintes, S., Brinkmann, B., Ebert, M. et al. Zeb2 is essential for Schwann cell differentiation, myelination and nerve repair. Nat Neurosci 19, 1050–1059 (2016). https://doi.org/10.1038/nn.4321
This article is cited by
Molecular Neurobiology (2023)
Relevance of biometals during neuronal differentiation and myelination: in vitro and in vivo studies
Neurochemical Research (2022)
Schwann cell plasticity regulates neuroblastic tumor cell differentiation via epidermal growth factor-like protein 8
Nature Communications (2021)