New directions in craniofacial morphogenesis

The vertebrate head is an extremely complicated structure: development of the head requires tissue-tissue interactions between derivates of all the germ layers and coordinated morphogenetic movements in three dimensions. In this review, we highlight a number of recent embryological studies, using chicken, frog, zebrafish and mouse, which have identified crucial signaling centers in the embryonic face. These studies demonstrate how small variations in growth factor signaling can lead to a diversity of phenotypic outcomes. We also discuss novel genetic studies, in human, mouse and zebrafish, which describe cell biological mechanisms fundamental to the growth and morphogenesis of the craniofacial skeleton. Together, these findings underscore the complex interactions leading to species-specific morphology. These and future studies will improve our understanding of the genetic and environmental influences underlying human craniofacial anomalies.     

Common mechanisms in development and disease: BMP signaling in craniofacial development

BMP signaling is one of the key pathways regulating craniofacial development. It is involved in the early patterning of the head, the development of cranial neural crest cells, and facial patterning. It regulates development of its mineralized structures, such as cranial bones, maxilla, mandible, palate, and teeth. Targeted mutations in the mouse have been instrumental to delineate the functional involvement of this signaling network in different aspects of craniofacial development. Gene polymorphisms and mutations in BMP pathway genes have been associated with various non-syndromic and syndromic human craniofacial malformations. The identification of intricate cellular interactions and underlying molecular pathways illustrate the importance of local fine-regulation of Bmp signaling to control proliferation, apoptosis, epithelial-mesenchymal interactions, and stem/progenitor differentiation during craniofacial development. Thus, BMP signaling contributes both to shape and functionality of our facial features. BMP signaling also regulates postnatal craniofacial growth and is associated with dental structures life-long. A more detailed understanding of BMP function in growth, homeostasis, and repair of postnatal craniofacial tissues will contribute to our ability to rationally manipulate this signaling network in the context of tissue engineering.     

Middle ear defects associated with the double knock out mutation of murine goosecoid and Msx1 genes.

A number of developmental regulatory genes, including homeobox genes, are dynamically expressed in the mammalian cephalic ectomesenchyme during craniofacial morphogenesis. Owing to the vast amount of gene knock out experiments, functions of such genes are now being revealed in the mammalian skeletal patterning process. The murine goosecoid (Gsc) and Msx1 genes are expressed during craniofacial development and each mutant mouse displays intriguing facial abnormalities including those of middle ear ossicles, suggesting that both genes play roles in spatial programming of craniofacial regions. In order to examine whether these genes could function in concert to direct particular craniofacial morphogenesis, double knock out mice were analyzed. The phenotype of the double mutant mice was restricted to the first arch derivatives and was apparently additive of the single gene mutant mice, implying region specific genetic interactions of these homeobox genes expressed in overlapping regions of middle ear forming ectomesenchyme. Our results also suggested that the patterning of distal portions of the malleus depends on the tympanic membrane, for which normal expressions of both the genes are prerequisite.     

A phenotype-driven ENU mutagenesis screen identifies novel alleles with functional roles in early mouse craniofacial development

Proper craniofacial development begins during gastrulation and requires the coordinated integration of each germ layer tissue (ectoderm, mesoderm, and endoderm) and its derivatives in concert with the precise regulation of cell proliferation, migration, and differentiation. Neural crest cells, which are derived from ectoderm, are a migratory progenitor cell population that generates most of the cartilage, bone, and connective tissue of the head and face. Neural crest cell development is regulated by a combination of intrinsic cell autonomous signals acquired during their formation, balanced with extrinsic signals from tissues with which the neural crest cells interact during their migration and differentiation. Although craniofacial anomalies are typically attributed to defects in neural crest cell development, the cause may be intrinsic or extrinsic. Therefore, we performed a phenotype-driven ENU mutagenesis screen in mice with the aim of identifying novel alleles in an unbiased manner, that are critically required for early craniofacial development. Here we describe 10 new mutant lines, which exhibit phenotypes affecting frontonasal and pharyngeal arch patterning, neural and vascular development as well as sensory organ morphogenesis. Interestingly, our data imply that neural crest cells and endothelial cells may employ similar developmental programs and be interdependent during early embryogenesis, which collectively is critical for normal craniofacial morphogenesis. Furthermore our novel mutants that model human conditions such as exencephaly, craniorachischisis, DiGeorge, and Velocardiofacial sydnromes could be very useful in furthering our understanding of the complexities of specific human diseases.     

Assessing Species-specific Contributions To Craniofacial Development Using Quail-duck Chimeras

The generation of chimeric embryos is a widespread and powerful approach to study cell fates, tissue interactions, and species-specific contributions to the histological and morphological development of vertebrate embryos. In particular, the use of chimeric embryos has established the importance of neural crest in directing the species-specific morphology of the craniofacial complex. The method described herein utilizes two avian species, duck and quail, with remarkably different craniofacial morphology. This method greatly facilitates the investigation of molecular and cellular regulation of species-specific pattern in the craniofacial complex. Experiments in quail and duck chimeric embryos have already revealed neural crest-mediated tissue interactions and cell-autonomous behaviors that regulate species-specific pattern in the craniofacial skeleton, musculature, and integument. The great diversity of neural crest derivatives suggests significant potential for future applications of the quail-duck chimeric system to understanding vertebrate development, disease, and evolution.     

Development of cell polarity in budding yeast

The development of cell polarity involves virtually every aspect of cell biology. Yeast are less complex than cells traditionally used for studies on cell polarity and are amendable to sophisticated genetic analysis. This has resulted in a growing number of molecular markers for yeast cell polarity and an increasingly well-defined progression of molecular events required for bud formation. Together, these factors provide a favorable context in which to understand how the interplay between a large number of processes can polarize a cell. Many genes required for morphogenesis have been identified, and genetic interactions provide evidence that the products of these genes function together. Studies on cell polarity development in S. cerevisiae have demonstrated a requirement for small GTP-binding proteins and have established functional relationships between temporally coincident events. With the continued identification and analysis of genes required for morphogenesis, and the pursuit of these studies on a cytological and biochemical level, studies on yeast will continue to contribute to our understanding of cell polarity development.     

Evolving maps in craniofacial development

The shaping of the vertebrate head results from highly dynamic integrated processes involving the growth and exchange of signals between the ectoderm, the endoderm, the mesoderm and Cephalic Neural Crest Cells (CNCCs). During embryonic development, these tissues change their shape and relative position rapidly and come transiently in contact with each other. Molecular signals exchanged in restricted regions of tissue interaction are crucial in providing positional identity to the mesenchymes which will form the different skeletal and muscular components of the head. Slight spatio-temporal modifications of these signalling maps can result in profound changes in craniofacial development and might have contributed to the evolution of facial diversity. Abnormal signalling patterns could also be at the origin of congenital craniofacial malformations. This review brings into perspective recent work on spatial and temporal aspects of facial morphogenesis with particular focus on the molecular mechanisms of jaw specification.     

Thyroid hormone and retinoic acid interact to regulate zebrafish craniofacial neural crest development

Craniofacial and ocular morphogenesis require proper regulation of cranial neural crest migration, proliferation, survival and differentiation. Although alterations in maternal thyroid hormone (TH) are associated with congenital craniofacial anomalies, the role of TH on the neural crest has not been previously described. Using zebrafish, we demonstrate that pharmacologic and genetic alterations in TH signaling disrupt cranial neural crest migration, proliferation, and survival, leading to craniofacial, extraocular muscle, and ocular developmental abnormalities. In the rostral cranial neural crest that gives rise to the periocular mesenchyme and the frontonasal process, retinoic acid (RA) rescued migratory defects induced by decreased TH signaling. In the caudal cranial neural crest, TH and RA had reciprocal effects on anterior and posterior pharyngeal arch development. The interactions between TH and RA signaling were partially mediated by the retinoid X receptor. We conclude that TH regulates both rostral and caudal cranial neural crest. Further, coordinated interactions of TH and RA are required for proper craniofacial and ocular development.     

Testing hypotheses about tinkering in the fossil record: the case of the human skull

Efforts to test hypotheses about small-scale shifts in development (tinkering) that can only be observed in the fossil record pose many challenges. Here we use the origin of modern human craniofacial form to explore a series of analytical steps with which to propose and test evolutionary developmental hypotheses about the basic modules of evolutionary change. Using factor and geometric morphometric analyses of craniofacial variation in modern humans, fossil hominids, and chimpanzee crania, we identify several key shifts in integration (defined as patterns of covariation that result from interactions between components of a system) among units of the cranium that underlie the unique shape of the modern human cranium. The results indicate that facial retraction in modern humans is largely a product of three derived changes: a relatively longer anterior cranial base, a more flexed cranial base angle, and a relatively shorter upper face. By applying the Atchley-Hall model of morphogenesis, we show that these shifts are most likely the result of changes in epigenetic interactions between the cranial base and both the brain and the face. Changes in the size of the skeletal precursors to these regions may also have played some role. This kind of phenotype-to-genotype approach is a useful and important complement to more standard genotype-to-phenotype approaches, and may help to identify candidate genes involved in the origin of modern human craniofacial form.     

Common genetic and environmental factors among craniofacial traits in Belgian nuclear families: Comparing skeletal and soft-tissue related phenotypes

The major objective of this study was to determine the possible effects of common genetic and environmental factors among 18 craniofacial anthropometric traits, with special attention to the differences between skeletal and soft-tissue related phenotypes. The studied sample consisted of 122 nuclear families living in Brussels and included 251 males and 258 females aged from 13 to 72 years. Univariate and bivariate quantitative genetic analyses were performed using a variance components procedure implemented in SOLAR software.All phenotypes were significantly influenced by additive genetic factors with heritability estimates ranging from 0.46 (nose height) to 0.72 (external biocular breadth). Sex, age and their interactions explained 7-46% of the total phenotypic variance of the traits. Bivariate analysis revealed that several traits share a common genetic and/or environmental basis while other traits show genetic and environmental independence from one another. More and greater genetic and environmental correlations were observed among skeletal phenotypes, than among soft-tissue traits and between both categories. Apart from the tissue composition, other characteristics of the craniofacial morphology such as the orientation (e.g. heights, breadths) have shown to be important factors in determining pleiotropy and common environmental effects between some pairs of traits. In conclusion, the results confirm that overall head configuration is largely determined by additive genetic effects, and that common genetic and environmental factors affecting craniofacial size and shape are stronger for the skeletal traits than for the soft-tissue traits.     

Branching morphogenesis of the lung: new molecular insights into an old problem

It has been known for decades that branching morphogenesis of the lung is mediated through reciprocal interactions between the epithelium and its underlying mesenchyme. In recent years, several key players, in particular members of the major signaling pathways that mediate this interaction, have been identified. Here, we review the genetic and molecular studies of these key components, which have provided a conceptual framework for understanding the interactions of these major signaling pathways in branching morphogenesis. The future challenge is to translate understanding of the signaling cascade into knowledge of the cellular responses, including cell proliferation, migration and differentiation, that lead to the stereotyped branching.*     

The role of sonic hedgehog in normal and abnormal craniofacial morphogenesis.

There is growing evidence that implicates a role for Sonic hedgehog (SHH) in morphogenesis of the craniofacial complex. Mutations in human and murine SHH cause midline patterning defects that are manifested in the head as holoprosencephaly and cyclopia. In addition, teratogens such as jervine, which inhibit the response of tissues to SHH, also produce cyclopia. Thus, the loss of SHH signaling during early stages of neural plate patterning has a profound influence of craniofacial morphogenesis. However, the severity of these defects precludes analyses of SHH function during later stages of craniofacial development. We have used an embryonic chick system to study the role of SHH during these later stages of craniofacial development. Using a combination of surgical and molecular experiments, we show here that SHH is essential for morphogenesis of the frontonasal and maxillary processes (FNP and MXPs), which give rise to the mid- and upper face. Transient loss of SHH signaling in the embryonic face inhibits growth of the primordia and results in defects analogous to hypotelorism and cleft lip/palate, characteristics of the mild forms of holoprosencephaly. In contrast, excess SHH leads to a mediolateral widening of the FNP and a widening between the eyes, a condition known as hypertelorism. In severe cases, this widening is accompanied by facial duplications. Collectively, these experiments demonstrate that SHH has multiple and profound effects on the entire spectrum of craniofacial development, and perturbations in SHH signaling are likely to underlie a number of human craniofacial anomalies.