Michael J. Dixon

The embryonic epidermis originates from a simple single-layered ectoderm that undergoes precise transcriptional and morphological transitions to generate a self-replenishing, multi-layered epidermis. A key early milestone is the emergence of the periderm; a transient, protective layer whose disruption underlies a series of devastating human congenital disorders. Despite its importance, the gene regulatory networks that drive periderm formation remain poorly defined. Here, we integrate single-cell multiome profiling with high-resolution spatial transcriptomics analysis in wild-type and transcription factor p63 mutant ( Trp63 ^-/- ) embryos to dissect the transcriptional and epigenetic programs driving epidermal lineage diversification. We resolve the cellular repertoire of early embryonic skin, define epithelial subpopulations and reconstruct the transcriptional dynamics underlying the periderm emergence. We show that p63 sits at the core of a multilayered enhancer-gene regulatory network, demonstrate how this network is perturbed in vivo , and functionally validate key targets using comparative cellular models. Together, these findings delineate the gene regulatory architecture of early skin development and establish a framework for understanding epithelial-associated congenital disorders.

Birth defects are the leading cause of infant mortality, and most inborn errors of development are multifactorial in origin resulting from complex gene-environment interactions. Definition of specific gene-environment interactions in the etiology and pathogenesis of congenital disorders is critically needed in the absence of genotype-phenotype correlation but is challenging. This is particularly true for congenital craniofacial anomalies, which account for approximately one-third of all birth defects, as they typically exhibit considerable inter- and intrafamilial variability. A classic example of this is Treacher Collins syndrome (TCS), which, although primarily caused by mutations in treacle ribosome biogenesis factor 1 ( TCOF1 ), is characterized by considerable variability in the severity of mandibulofacial dysostosis. Here, we describe the genetic and environmental factors with converging effects that mechanistically contribute to the etiology and pathogenesis of craniofacial variation in this rare congenital disorder. We discovered in Tcof1 ^+/– mouse models of TCS that the combination of different endogenous levels of Tcof1 (also known as treacle) protein and ROS within distinct genetic backgrounds correlated with TCS phenotype severity. Furthermore, geometric morphometric analyses revealed that genotype largely determines the craniofacial shape but that redox status determines the size of individual bones. Taken together, our results highlight the roles of ROS and genomic instability in modulating the variability and phenotype severity of craniofacial anomalies.

This paper provides critical reflection on work and progress to embed EDI in our research and innovation workforce, practice and culture at the University of Manchester. Our university aim is to take an intersectional and holistic approach to educate, engage, empower and support our staff and student community at all levels to prioritise EDI. To embed EDI in research and promote collective responsibility to help shape a fairer, inclusive research culture the University launched the ‘Inclusive Research Transformation Programme’. This programme included developing inclusive research mindsets and building inclusive leadership capacity (from UG to senior research leader); a University EDI award scheme which catalysed innovative local and national researcher led EDI initiatives; and targeted funding schemes to help address gender, ethnicity, and disability inequities within our research career pipeline. Three schemes were developed, an UG EDI summer placement scheme, an early career research staff EDI fellowship underpinned by inclusive advocacy and an established academic returners scheme. Funding AcknowledgementThis work was funded by University of Manchester and Wellcome Institutional Strategic Support Fund (204796/Z/16/Z ISSF - Wellcome ISSF 3 – EDI).

Clefts of the lip and palate (CLP), the major causes of congenital facial malformation globally, result from failure of fusion of the facial processes during embryogenesis. With a prevalence of 1 in 500–2500 live births, CLP causes major morbidity throughout life as a result of problems with facial appearance, feeding, speaking, obstructive apnoea, hearing and social adjustment and requires complex, multi‐disciplinary care at considerable cost to healthcare systems worldwide. Long‐term outcomes for affected individuals include increased mortality compared with their unaffected siblings. The frequent occurrence and major healthcare burden imposed by CLP highlight the importance of dissecting the molecular mechanisms driving facial development. Identification of the genetic mutations underlying syndromic forms of CLP, where CLP occurs in association with non‐cleft clinical features, allied to developmental studies using appropriate animal models is central to our understanding of the molecular events underlying development of the lip and palate and, ultimately, how these are disturbed in CLP.

Cleft palate is a common birth defect that frequently occurs in human congenital malformations caused by mutations in components of the Sonic Hedgehog (SHH) signaling cascade. Shh is expressed in dynamic, spatiotemporal domains within epithelial rugae and plays a key role in driving epithelial-mesenchymal interactions that are central to development of the secondary palate. However, the gene regulatory networks downstream of Hedgehog (Hh) signaling are incompletely characterized. Here, we show that ectopic Hh signaling in the palatal mesenchyme disrupts oral-nasal patterning of the neural crest cell–derived ectomesenchyme of the palatal shelves, leading to defective palatine bone formation and fully penetrant cleft palate. We show that a series of Fox transcription factors, including the novel direct target Foxl1, function downstream of Hh signaling in the secondary palate. Furthermore, we demonstrate that Wnt/bone morphogenetic protein (BMP) antagonists, in particular Sostdc1, are positively regulated by Hh signaling, concomitant with downregulation of key regulators of osteogenesis and BMP signaling effectors. Our data demonstrate that ectopic Hh-Smo signaling downregulates Wnt/BMP pathways, at least in part by upregulating Sostdc1, resulting in cleft palate and defective osteogenesis.

Cleft palate is a common congenital disorder that affects up to 1 in 2500 live births and results in considerable morbidity to affected individuals and their families. The aetiology of cleft palate is complex with both genetic and environmental factors implicated. Mutations in the transcription factor p63 are one of the major individual causes of cleft palate; however, the gene regulatory networks in which p63 functions remain only partially characterized. Our findings demonstrate that p63 functions as an essential regulatory molecule in the spatio-temporal control of palatal epithelial cell fate to ensure appropriate fusion of the palatal shelves. Initially, p63 induces periderm formation and controls its subsequent maintenance to prevent premature adhesion between adhesion-competent, intra-oral epithelia. Subsequently, TGFβ3-induced down-regulation of p63 in the medial edge epithelia of the palatal shelves is a pre-requisite for palatal fusion by facilitating periderm migration from, and reducing the proliferative potential of, the midline epithelial seam thereby preventing cleft palate. Cleft palate is a serious congenital condition which affects approximately 1 in every 2500 births. Cleft palate occurs when the palatal shelves fail to grow, adhere or fuse during development. Mutations in the gene encoding the transcription factor p63 result in cleft palate in humans and mice. However, the role of p63 and how it controls the network of genes to regulate palate development is not well understood.In this study, we demonstrate that p63 controls the spatio-temporal regulation of palatal epithelial cell fate to ensure appropriate palatal adhesion: p63 induces the formation of a flattened layer of epithelial (periderm) cells and controls its subsequent maintenance. We also demonstrate that TGFβ3-induced, down-regulation of p63 in the medial edge epithelial cells, through which the palatal shelves adhere and fuse, controls Jag2-induced periderm migration to the oral and nasal epithelial triangles. In addition, p63 plays a central role in maintaining the proliferative potential of the basal layer of the medial edge epithelia. Our study provides significant new insights into the mechanisms that regulate development of the palate by establishing the role of p63 in governing the fate of the midline epithelial cells.

Craniofacial anomalies account for approximately one‐third of all congenital birth defects reflecting the complexity of head and facial development. Craniofacial development is dependent upon a multipotent, migratory population of neural crest cells, which generate most of the bone and cartilage of the head and face. In this review, we discuss advances in our understanding of the pathogenesis of a specific array of craniofacial anomalies, termed facial dysostoses, which can be subdivided into mandibulofacial dysostosis, which present with craniofacial defects only, and acrofacial dysostosis, which encompasses both craniofacial and limb anomalies. In particular, we focus on Treacher Collins syndrome, Acrofacial Dysostosis‐Cincinnati Type as well as Nager and Miller syndromes, and animal models that provide new insights into the molecular and cellular basis of these congenital syndromes. We emphasize the etiologic and pathogenetic similarities between these birth defects, specifically their unique deficiencies in global processes including ribosome biogenesis, DNA damage repair, and pre‐mRNA splicing, all of which affect neural crest cell development and result in similar tissue‐specific defects. WIREs Dev Biol 2017, 6:e263. doi: 10.1002/wdev.263This article is categorized under: Signaling Pathways > Cell Fate Signaling Birth Defects > Craniofacial and Nervous System Anomalies

Appropriate development of stratified, squamous, keratinizing epithelia, such as the epidermis and oral epithelia, generates an outer protective permeability barrier that prevents water loss, entry of toxins, and microbial invasion. During embryogenesis, the immature ectoderm initially consists of a single layer of undifferentiated, cuboidal epithelial cells that stratifies to produce an outer layer of flattened periderm cells of unknown function. Here, we determined that periderm cells form in a distinct pattern early in embryogenesis, exhibit highly polarized expression of adhesion complexes, and are shed from the outer surface of the embryo late in development. Mice carrying loss-of-function mutations in the genes encoding IFN regulatory factor 6 (IRF6), IκB kinase-α (IKKα), and stratifin (SFN) exhibit abnormal epidermal development, and we determined that mutant animals exhibit dysfunctional periderm formation, resulting in abnormal intracellular adhesions. Furthermore, tissue from a fetus with cocoon syndrome, a lethal disorder that results from a nonsense mutation in IKKA, revealed an absence of periderm. Together, these data indicate that periderm plays a transient but fundamental role during embryogenesis by acting as a protective barrier that prevents pathological adhesion between immature, adhesion-competent epithelia. Furthermore, this study suggests that failure of periderm formation underlies a series of devastating birth defects, including popliteal pterygium syndrome, cocoon syndrome, and Bartsocas-Papas syndrome.

Inherited diseases caused by genetic mutations can arise due to loss of protein function. Alternatively, mutated proteins may mis-fold, impairing endoplasmic reticulum (ER) trafficking, causing ER stress and triggering the unfolded protein response (UPR). The UPR attempts to restore proteostasis but if unsuccessful drives affected cells towards apoptosis. Previously, we reported that in mice, the p.Tyr64His mutation in the enamel extracellular matrix (EEM) protein amelogenin disrupts the secretory pathway in the enamel-forming ameloblasts, resulting in eruption of malformed tooth enamel that phenocopies human amelogenesis imperfecta (AI). Defective amelogenin post-secretory self-assembly and processing within the developing EEM has been suggested to underlie the pathogenesis of X chromosome-linked AI. Here, we challenge this concept by showing that AI pathogenesis associated with the p.Tyr64His amelogenin mutation involves ameloblast apoptosis induced by ER stress. Furthermore, we show that 4-phenylbutyrate can rescue the enamel phenotype in affected female mice by promoting cell survival over apoptosis such that they are able to complete enamel formation despite the presence of the mutation, offering a potential therapeutic option for patients with this form of AI and emphasizing the importance of ER stress in the pathogenesis of this inherited conformational disease.