HDAC1 and HDAC2 control the specification of neural crest cells into peripheral glia.
ABSTRACT: Schwann cells, the myelinating glia of the peripheral nervous system (PNS), originate from multipotent neural crest cells that also give rise to other cells, including neurons, melanocytes, chondrocytes, and smooth muscle cells. The transcription factor Sox10 is required for peripheral glia specification. However, all neural crest cells express Sox10 and the mechanisms directing neural crest cells into a specific lineage are poorly understood. We show here that histone deacetylases 1 and 2 (HDAC1/2) are essential for the specification of neural crest cells into Schwann cell precursors and satellite glia, which express the early determinants of their lineage myelin protein zero (P0) and/or fatty acid binding protein 7 (Fabp7). In neural crest cells, HDAC1/2 induced expression of the transcription factor Pax3 by binding and activating the Pax3 promoter. In turn, Pax3 was required to maintain high Sox10 levels and to trigger expression of Fabp7. In addition, HDAC1/2 were bound to the P0 promoter and activated P0 transcription. Consistently, in vivo genetic deletion of HDAC1/2 in mouse neural crest cells led to strongly decreased Sox10 expression, no detectable Pax3, virtually no satellite glia, and no Schwann cell precursors in dorsal root ganglia and peripheral nerves. Similarly, in vivo ablation of Pax3 in the mouse neural crest resulted in strongly reduced expression of Sox10 and Fabp7. Therefore, by controlling the expression of Pax3 and the concerted action of Pax3 and Sox10 on their target genes, HDAC1/2 direct the specification of neural crest cells into peripheral glia.
Project description:Sox proteins are characterized by possession of a DNA-binding domain with similarity to the high-mobility group domain of the sex determining factor SRY. Here, we report on Sox10, a novel protein with predominant expression in glial cells of the nervous system. During development Sox10 first appeared in the forming neural crest and continued to be expressed as these cells contributed to the forming PNS and finally differentiated into Schwann cells. In the CNS, Sox10 transcripts were originally confined to glial precursors and later detected in oligodendrocytes of the adult brain. Functional studies failed to reveal autonomous transcriptional activity for Sox10. Instead, Sox10 functioned synergistically with the POU domain protein Tst-1/Oct6/SCIP with which it is coexpressed during certain stages of Schwann cell development. Synergy depended on binding to adjacent sites in target promoters, was mediated by the N-terminal regions of both proteins, and could not be observed between Sox10 and several other POU domain proteins. Interestingly, Sox10 also modulated the function of Pax3 and Krox-20, two other transcription factors involved in Schwann cell development. We propose a role for Sox10 in conferring cell specificity to the function of other transcription factors in developing and mature glia.
Project description:The transcription factor Sox5 has previously been shown in chicken to be expressed in early neural crest cells and neural crest-derived peripheral glia. Here, we show in mouse that Sox5 expression also continues after neural crest specification in the melanocyte lineage. Despite its continued expression, Sox5 has little impact on melanocyte development on its own as generation of melanoblasts and melanocytes is unaltered in Sox5-deficient mice. Loss of Sox5, however, partially rescued the strongly reduced melanoblast generation and marker gene expression in Sox10 heterozygous mice arguing that Sox5 functions in the melanocyte lineage by modulating Sox10 activity. This modulatory activity involved Sox5 binding and recruitment of CtBP2 and HDAC1 to the regulatory regions of melanocytic Sox10 target genes and direct inhibition of Sox10-dependent promoter activation. Both binding site competition and recruitment of corepressors thus help Sox5 to modulate the activity of Sox10 in the melanocyte lineage.
Project description:Hirschsprung disease and Waardenburg syndrome are human genetic diseases characterized by distinct neural crest defects. Patients with Hirschsprung disease suffer from gastrointestinal motility disorders, whereas Waardenburg syndrome consists of defective melanocyte function, deafness, and craniofacial abnormalities. Mutations responsible for Hirschsprung disease and Waardenburg syndrome have been identified, and some patients have been described with characteristics of both disorders. Here, we demonstrate that PAX3, which is often mutated in Waardenburg syndrome, is required for normal enteric ganglia formation. Pax3 can bind to and activate expression of the c-RET gene, which is often mutated in Hirschsprung disease. Pax3 functions with Sox10 to activate transcription of c-RET, and SOX10 mutations result in Waardenburg-Hirschsprung syndrome. Thus, Pax3, Sox10, and c-Ret are components of a neural crest development pathway, and interruption of this pathway at various stages results in neural crest-related human genetic syndromes.
Project description:The neural crest is a multipotent population of cells that originates a variety of cell types. Many animal models are used to study neural crest induction, migration and differentiation, with amphibians and birds being the most widely used systems. A major technological advance to study neural crest development in mouse, chick and zebrafish has been the generation of transgenic animals in which neural crest specific enhancers/promoters drive the expression of either fluorescent proteins for use as lineage tracers, or modified genes for use in functional studies. Unfortunately, no such transgenic animals currently exist for the amphibians Xenopus laevis and tropicalis, key model systems for studying neural crest development. Here we describe the generation and characterization of two transgenic Xenopus laevis lines, Pax3-GFP and Sox10-GFP, in which GFP is expressed in the pre-migratory and migratory neural crest, respectively. We show that Pax3-GFP could be a powerful tool to study neural crest induction, whereas Sox10-GFP could be used in the study of neural crest migration in living embryos.
Project description:Schwann cells (SCs) arise from neural crest cells (NCCs) that first give rise to SC precursors (SCPs), followed by immature SCs, pro-myelinating SCs, and finally, non-myelinating or myelinating SCs. After nerve injury, mature SCs 'de-differentiate', downregulating their myelination program while transiently re-activating early glial lineage genes. To better understand molecular parallels between developing and de-differentiated SCs, we characterized the expression profiles of a panel of 12 transcription factors from the onset of NCC migration through postnatal stages, as well as after acute nerve injury. Using Sox10 as a pan-glial marker in co-expression studies, the earliest transcription factors expressed in E9.0 Sox10+ NCCs were Sox9, Pax3, AP2? and Nfatc4. E10.5 Sox10+ NCCs coalescing in the dorsal root ganglia differed slightly, expressing Sox9, Pax3, AP2? and Etv5. E12.5 SCPs continued to express Sox10, Sox9, AP2? and Pax3, as well as initiating Sox2 and Egr1 expression. E14.5 immature SCs were similar to SCPs, except that they lost Pax3 expression. By E18.5, AP2?, Sox2 and Egr1 expression was turned off in the nerve, while Jun, Oct6 and Yy1 expression was initiated in pro-myelinating Sox9+/Sox10+ SCs. Early postnatal and adult SCs continued to express Sox9, Jun, Oct6 and Yy1 and initiated Nfatc4 and Egr2 expression. Notably, at all stages, expression of each marker was observed only in a subset of Sox10+ SCs, highlighting the heterogeneity of the SC pool. Following acute nerve injury, Egr1, Jun, Oct6, and Sox2 expression was upregulated, Egr2 expression was downregulated, while Sox9, Yy1, and Nfatc4 expression was maintained at similar frequencies. Notably, de-differentiated SCs in the injured nerve did not display a transcription factor profile corresponding to a specific stage in the SC lineage. Taken together, we demonstrate that uninjured and injured SCs are heterogeneous and distinct from one another, and de-differentiation recapitulates transcriptional aspects of several different embryonic stages.
Project description:CHD7 mutations are implicated in a majority of cases of the congenital disorder, CHARGE syndrome. CHARGE, an autosomal dominant syndrome, is known to affect multiple tissues including eye, heart, ear, craniofacial nerves and skeleton and genital organs. Using a morpholino-antisense-oligonucleotide-based zebrafish model for CHARGE syndrome, we uncover a complex spectrum of abnormalities in the neural crest and the crest-derived cell types. We report for the first time, defects in myelinating Schwann cells, enteric neurons and pigment cells in a CHARGE model. We also observe defects in the specification of peripheral neurons and the craniofacial skeleton as previously reported. Chd7 morphants have impaired migration of neural crest cells and deregulation of sox10 expression from the early stages. Knocking down Sox10 in the zebrafish CHARGE model rescued the defects in Schwann cells and craniofacial cartilage. Our zebrafish CHARGE model thus reveals important regulatory roles for Chd7 at multiple points of neural crest development viz., migration, fate choice and differentiation and we suggest that sox10 deregulation is an important driver of the neural crest-derived aspects of Chd7 dependent CHARGE syndrome.
Project description:Systemic loss-of-function studies have demonstrated that Pax3 transcription factor expression is essential for dorsal neural tube, early neural crest and muscle cell lineage morphogenesis. Cardiac neural crest cells participate in both remodeling of the pharyngeal arch arteries and outflow tract septation during heart development, but the lineage specific role of Pax3 in neural crest function has not yet been determined. To gain insight into the requirement of Pax3 within the neural crest, we conditionally deleted Pax3 in both the premigratory and migratory neural crest populations via Wnt1-Cre and Ap2?-Cre and via P0-Cre in only the migratory neural crest, and compared these phenotypes to the pulmonary atresia phenotype observed following the systemic loss of Pax3. Surprisingly, using Wnt1-Cre deletion there are no resultant heart defects despite the loss of Pax3 from the premigratory and migratory neural crest. In contrast, earlier premigratory and migratory Ap2?-Cre mediated deletion resulted in double outlet right ventricle alignment heart defects. In order to assess the tissue-specific contribution of neural crest to heart development, genetic ablation of neural crest lineage using a Wnt1-Cre-activated diphtheria toxin fragment-A cell-killing system was employed. Significantly, ablation of Wnt1-Cre-expressing neural crest cells resulted in fully penetrant persistent truncus arteriosus malformations. Combined, the data show that Pax3 is essential for early neural crest progenitor formation, but is not required for subsequent cardiac neural crest progeny morphogenesis involving their migration to the heart or septation of the outflow tract.
Project description:BACKGROUND: While several mouse strains have recently been developed for tracing neural crest or oligodendrocyte lineages, each strain has inherent limitations. The connection between human SOX10 mutations and neural crest cell pathogenesis led us to focus on the Sox10 gene, which is critical for neural crest development. We generated Sox10-Venus BAC transgenic mice to monitor Sox10 expression in both normal development and in pathological processes. RESULTS: Tissue fluorescence distinguished neural crest progeny cells and oligodendrocytes in the Sox10-Venus mouse embryo. Immunohistochemical analysis confirmed that Venus expression was restricted to cells expressing endogenous Sox10. Time-lapse imaging of various tissues in Sox10-Venus mice demonstrated that Venus expression could be visualized at the single-cell level in vivo due to the intense, focused Venus fluorescence. In the adult Sox10-Venus mouse, several types of mature and immature oligodendrocytes along with Schwann cells were clearly labeled with Venus, both before and after spinal cord injury. CONCLUSIONS: In the newly-developed Sox10-Venus transgenic mouse, Venus fluorescence faithfully mirrors endogenous Sox10 expression and allows for in vivo imaging of live cells at the single-cell level. This Sox10-Venus mouse will thus be a useful tool for studying neural crest cells or oligodendrocytes, both in development and in pathological processes.
Project description:In the current study, we examined the role of Ezh2 as an epigenetic modifier for the enteric neural crest cell development through H3K27me3. Ezh2 conditional null mice were viable up to birth, but died within the first hour of life. In addition to craniofacial defects, Ezh2 conditional null mice displayed reduced number of ganglion cells in the enteric nervous system. RT-PCR and ChIP assays indicated aberrant up-regulation of Zic1, Pax3, and Sox10 and loss of H3K27me3 marks in the promoter regions of these genes in the myenteric plexus. Overall, these results suggest that Ezh2 is an important epigenetic modifier for the enteric neural crest cell development through repression of Zic1, Pax3, and Sox10.
Project description:We show that highly pure populations of human Schwann cells can be derived rapidly and in a straightforward way, without the need for genetic manipulation, from human epidermal neural crest stem cells [hEPI-NCSC(s)] present in the bulge of hair follicles. These human Schwann cells promise to be a useful tool for cell-based therapies, disease modelling and drug discovery. Schwann cells are glia that support axons of peripheral nerves and are direct descendants of the embryonic neural crest. Peripheral nerves are damaged in various conditions, including through trauma or tumour-related surgery, and Schwann cells are required for their repair and regeneration. Schwann cells also promise to be useful for treating spinal cord injuries. Ex vivo expansion of hEPI-NCSC isolated from hair bulge explants, manipulating the WNT, sonic hedgehog and TGFβ signalling pathways, and exposure of the cells to pertinent growth factors led to the expression of the Schwann cell markers SOX10, KROX20 (EGR2), p75NTR (NGFR), MBP and S100B by day 4 in virtually all cells, and maturation was completed by 2 weeks of differentiation. Gene expression profiling demonstrated expression of transcripts for neurotrophic and angiogenic factors, as well as JUN, all of which are essential for nerve regeneration. Co-culture of hEPI-NCSC-derived human Schwann cells with rodent dorsal root ganglia showed interaction of the Schwann cells with axons, providing evidence of Schwann cell functionality. We conclude that hEPI-NCSCs are a biologically relevant source for generating large and highly pure populations of human Schwann cells.