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The society of craniofacial genetics and developmental biology 36th annual meeting

2014; Wiley; Volume: 164; Issue: 8 Linguagem: Inglês

10.1002/ajmg.a.36565

1552-4833

Matthew P. Harris, Patricia L. Purcell,

Craniofacial Disorders and Treatments

There is nothing like October baseball in Boston. Autumn in New England already brings with it the feeling of change with the progression of the seasons and influx of students, but this year the atmosphere around the Fens and Longwood Medical Area encircling Fenway Park, this feeling was palpable. This was the setting of the 36th Annual Meeting of the Society of Craniofacial Genetics and Developmental Biology (SCGDB) held at Boston Children's Hospital and Harvard Medical School. As the SCGDB meeting is routinely scheduled to co-occur with the annual meeting of the American Society for Human Genetics, the SCGDB annual meeting brought together international investigators of diverse backgrounds with common interest in understanding the regulation of craniofacial development and the causes of disease. Plenary talks and posters were presented by experts in diverse fields of science with common theme in the genetic mechanisms underlying variation in form and function of the head. The mission of the Society of Craniofacial Genetics and Developmental Biology centers on cross discipline communication between researchers of diverse fields to foster a broader understanding of the mechanisms underlying craniofacial growth, variation, and disease (http://craniofacialgenetics.org). A major emphasis in this goal is to integrate and encourage young investigators within the community. Accordingly, a large number of speakers in the meeting were postdoctoral researchers presenting their new work. Here, we present a brief overview of the 2013 meeting followed by abstracts of the posters presented at the meeting. The 37th annual meeting of the SCGDB will be held in San Diego, CA on Saturday, October 18, 2014. The increase use of genomic tools has revolutionized systematic approaches for understanding genetic, and epigenetic regulation of development and disease. The beginning session of the meeting highlighted talks describing new methods in clinical and basic research to define changes within regulatory domains as well as sequences encoding proteins influencing variation in skull development. The first speaker, Justin Cotney (Noonan lab, Yale University) described his work on genome wide identification of enhancers regulating development of the face and their association with clefting in patients. In a comparative approach, Nicolas Rohner (Tabin lab, Harvard Medical School) described his work in use of comparative genomics in primates to identify essential regulatory domains in the evolution of the human skull. Reintroduction of differentially retained enhancers stemming from his analysis in the developing mouse resulted in clues at the structures regulated by the underlying differentially retained gene expression. Following, Denise Liberton (University of Calgary) expanded on how variation within populations can be used to identify loci that are currently under selection in skull form. The combination of approaches from evolutionary signatures of macroevolutionary changes to population level variance of developmental regulators nicely integrate to probe the regulation associated with changes in craniofacial form. Although changes in cis-regulatory domains are essential in shaping the form and function of the skull, mutations affecting coding sequence are a common cause of developmental abnormalities and disease. Andrew Wilkie (Oxford University) detailed in this session his group's work on whole exome sequencing of patients to identify causative mutations underlying craniosynostosis. He detailed, that through use of a large cohort of patients, the use of whole exome sequencing can efficiently identify altered genetic variants underlying premature suture fusion. The power of these genomic approaches requires functional validation in models to systematically assess function of these genetic variants and their function in the development of the skull. He discussed his labs work on assessing the role of several variants in craniofacial development and effect on suture patency. One of the trends observed across the wide variety of research presented was the use of comparative analysis of gene function and discovery in different model organisms to address fundamental questions in craniofacial development. The majority of oral and poster presentations at this meeting used mouse, zebrafish, chicken, as well as Xenopus to address questions of changes in gene function and or developmental mechanisms. However, a growing number of studies also highlighted the use of less conventional species to identify critical mechanisms underlying craniofacial variation. The second section focused on comparative analysis of diverse organisms to define the mechanisms underlying variation of the skull. Elaine Ostrander (NIH NHGRI) presented the first plenary lecture on her work that uses diversity among dog breeds to help define the fundamental genetic basis of shape change in the skull. Elaine described both published and new work on defining unexpected genetic regulators of facial form though analysis of quantitative variation between different dog breeds. The function of genes identified in Dr. Ostrander's analysis were assessed by zebrafish and mouse models exemplifying use of comparative analysis to verify function of particular variants in craniofacial development. Kara Powder (Albertson Lab, University of Massachusetts Amherst) followed discussing a similar genetic approach however looking at species that, unlike dogs and the intense selection that shaped their skulls, showed selective change in facial form due to natural selection. She described the identification of mutations in the limb bud homeobox gene (lbx) as underlying a component of the change in jaw shape observed in speciation among different fish species of the family Cichlidae (Cichlids). The function of lbx variants found in the cichlids was tested in the zebrafish as well as Xenopus demonstrating a role for this gene on neural crest migration. Richard Schneider (University of California, San Francisco) and James Hanken (Harvard University) presented work from their laboratories defining properties of constraint and diversity in development of the vertebrate skull through a comparative analysis between closely related but divergent species. Rich Schneider defined mechanisms for shape change within the avian jaw by capitalizing on differences in development of the jaw skeleton between chicken, duck and emu and the ability to do cross species transplants early in development. Similarly, James Hanken discussed the utility of lineage tracing of neural crest in amphibians to identify homology as well as the potential/constraint of different cellular lineages to establish the shape of the skull. The use of model organisms in genetic analysis has been bolstered by the ability to perform mutagenesis screens for unbiased identification of gene function in development. Katrin Henke (Harris Lab, Boston Children's Hospital, Harvard Medical School) presented her recent work using the zebrafish as a model for craniofacial development and disease, emphasizing the analysis of late developmental events in growth and differentiation that lead to formation of the form of the skull. She described results of her large-scale screen for genes affecting form of the zebrafish skull, in particular those mutants that lead to changes in suture patency and patterning. The third session of the meeting focused on uncovering cellular and molecular mechanisms of craniofacial development. Richard Maas (Bringham and Women's Hospital; Harvard Medical School) presented the second plenary lecture detailing his work on the developmental mechanisms involved in orofacial clefting. The use of clinical genetics defined the unique role of SPECC1L in regulating the cytoskeleton affecting the development of the cartilage of the frontonasal bones. They have explored the function of SPECC1L and other novel clefting genes in regulating pattern in development using analysis of patterning in Drosophila, and functional analysis of patterning of the larval cranial cartilages in zebrafish through collaboration with lab of Dr. Eric Lao (Mass General Hospital, Harvard Medical School). The second talk of this session was given by Lisa L. Sandell (University of Louisville) who discussed the work of her lab in defining interactions between tissues of neural crest and otic vesicle derived neurons in the development of the inner ear. Using dynamic imaging in the embryonic mouse and experimental perturbation in the chick, she described the dependence of migratory neural crest cells in patterning of the cochleovestibular nerve. Following, Erika Kague (Fischer Lab, University of Pennsylvania) presented her work on the role of changes in rates of osteogenic differentiation on shape change and patterning of the skull. Using a zebrafish model defective in sp7, she observed that osteogenic differentiation was delayed and led to specific changes in shape and interaction of the skull roof and frontonasal aspects of the adult skull in addition to defects of tooth differentiation and replacement. Then, James Martin (Baylor College of Medicine) detailed the role of microRNA in the regulation of frontonasal development and clefting. Mouse deficient in the miR-17-92 cluster demonstrated cleft lip and mandibular hypoplasia in part through regulation of Tbx1 and Tbx3 expression during development of the jaw. He outlined evidence that miR-17-92 is regulated by Ap-2α and this microRNA is an essential regulator of the development of the mouth and jaw. Lastly, Katherine Fantauzzo (Soriano lab, Icahn School of Medicine at Mount Sinai) detailed signaling mechanisms essential for normal palate development, centering on PI3K and PDGFRα mediated signaling. Looking at PDGFRα mutant mice in which binding of PDGFRα to PI3K is disrupted, she analyzed differentially phosphorylated substrates of Akt and identified proteins that regulate cell survival and proliferation. Consistent with these findings, they found interaction of PDGFRα downstream effectors and deficiencies in the p53 gene, leading to compound effects on palate development. The final session of the oral presentations centered on genetic analysis of patients and approaches integrating these data into novel therapeutic approaches. Richard Sherwood (Wright State University) presented recent data stemming from analysis of normal variation of teeth and jaws among the Jirels of the Jiri valley in Eastern Nepal. This population is interesting as it contains a robust pedigree allowing for heritability of different orofacial traits to be ascertained. Morphometric analysis of variation in this population showed significant association of several loci to morphological variance of the palate and dentition. The discussion was followed by presentation by Myrian Peyard (Karolinska Institute, Sweeden) of the discovery of novel mutations in GRAINYHEAD 3 underlying Van der Woude syndrome and cleft palate. Analysis in both zebrafish and mouse models suggest that the mutations may impart a dominant negative effect that results in abnormal oral periderm development contributing to clefting. The following two talks centered on the role of variation in levels of signaling during development and the implications for understanding variant expressivity of phenotypes of the skull. Jennifer Fish (Marcucio lab, University of California, San Francisco) described elegant work on reduction of Fgf8 signaling on facial variation in the mouse. Using an allelic series of different mutants with effects on Fgf8 function she described the appearance of variation in expression as well as skeletal form once threshold levels of fgf8 were surpassed by accumulation of variant alleles. Nandini Singh (Richtsmeier lab, Pennsylvania State University) followed with a discussion of her work, quantitatively assessing the effects of a Sonic hedgehog (Shh) agonist, SAG, on craniofacial morphology as a step towards the potential use of SHH-agonists as a pharmaceutical therapy for patients with Down syndrome (DS). They found that Shh agonist-based treatments for DS should proceed with caution due to the severe dysmorphologies it causes to the craniofacial skeleton. They additionally found that dosage effects of the SAG-agonist play an important role in variability in craniofacial morphology. Dr. Priscilla Chen, friend of many and Scientific Research Officer of the National Institute of Dental and Craniofacial Research SBDD study section, died unexpectedly on October 11, 2013. Her commitment to improve the quality of research through careful and fair review touched the lives of countless scientists. The keynote lecture was named in her honor in memory of her lasting impact on the scientific community and the field. Dr. Bjorn Olsen (Harvard school of Dental Medicine) closed the oral presentations of the meeting with the Priscilla Chen Keynote lecture. Dr. Olsen discussed his work on cellular signaling mechanisms underlying vascular anomalies in children specifically related with the formation of vascular tumors. His work points to a role of altered vascular endothelial growth factor receptor signaling as a primary marker of hemangioma endothelial cells. He detailed the regulation of VEGFR1 expression by a receptor complex involving B1-INTEGRIN, VEGFR2, and TEM8/anthrax toxin receptor 1 (ANTXR1). Patients with infantile hemangioma harbor germline heterozygous missense mutations in genes encoding VEGFR2 and TEM8/ANTXR1. These variants cause increased interaction of the receptor complex and a decrease in integrin/NFAT signaling. Interestingly, TEM8/ANTXR1 homozygous null mutations have been associated with GAPO syndrome leading to a complex disorder including craniofacial defects of plagiocephaly, pseudoanodontia and optic atrophy. The syndrome is associated with increased extracellular material and venous malformations. This finding ties the action of TEM8/ANTXR1, and its regulation of VEGFR function to regulation of craniofacial morphogenesis during development. Polr1c and Polr1d knock-out mice as new genetic models for Treacher Collins Syndrome Treacher Collins syndrome (TCS) is a congenital disorder characterized by craniofacial deformities including external and middle ear defects, cleft palate, and hypoplasia of facial bones. TCOF1 underlies TCS in humans and a mouse model generated previously exhibits phenotypes similar to human TCS caused by a decrease in cranial neural crest cells (CNCC) that give rise to the majority of bone in the head and face. Tcof1 encodes the nucleolar phosphoprotein Treacle which functions in ribosomal DNA (rDNA) transcription. Tcof1+/− mice exhibit death of CNCC precursors due to a reduction in mature ribosomes. Recently, two more genes involved in rDNA transcription have been identified in TCS patients: POLR1C and POLR1D. We sought to investigate the role of these genes in the pathology of TCS by knocking out each gene in the mouse. Deletion of Polr1c specifically in the neural crest causes a reduction of migrating NCCs that has devastating effects in the development of the facial prominences including the 1st and 2nd pharyngeal arches and the frontonasal region. These embryos die at midgestation but temporal excision of Polr1c uncovered later craniofacial deformities that resemble human TCS. In addition, by generating compound mutants we were able to uncover genetic interactions between Polr1c, Polr1d, and Tcof1. The phenotypes of the new knock-out mice resemble the ones observed in the Tcof1 mice and in human TCS. Therefore these mouse models cannot only help us dissect out the etiology of this syndrome but they also provide avenues for the potential prevention this neurocristopathy disorder. Wnt-signaling underlies key aspects of cichlid craniofacial diversity East-African cichlids are unquestionably one of the most phenotypically diverse vertebrate lineages on the planet. Key to their success has been the evolution of divergent feeding morphologies, making them an ideal system in which to examine the genetic basis of craniofacial variation. Here we show that variation in Wnt/β-catenin-signaling underlies key aspects of cichlid craniofacial diversity. Specifically, we show that increased Wnt-signaling is associated with the development of a novel, lineage-specific craniofacial morphology, and that experimental modulation of the Wnt pathway recapitulates natural variation in craniofacial form. We suggest that expanded Wnt expression early in development serves to “lock” an adult phenotype into place within the phenotypically derived cichlid lineage, and that this likely occurs through early and/or expanded differentiation of bone forming cells. Notably, we show that the canalized phenotype in cichlids is associated with an increased sensitivity to molecular perturbation and a concomitant reduction in developmental plasticity, two features that should, in theory, limit evolutionary potential. In all, our data implicate the Wnt pathway as an important mediator of craniofacial form and offer new insights into how developmental systems can evolve to both promote and constrain evolutionary change. Wdr43, a ribosome biogenesis factor, regulates zebrafish neural crest development The neural Crest (NC), a population of cells originating from the dorsal side of the embryonic neural tube, contributes a significant portion of craniofacial skeleton. Defects in NC specification, migration, and differentiation lead to craniofacial malformations. In recent years, it has become evident that a number of genes regulating ribosome biogenesis also contribute to craniofacial development. Through an ENU chemical mutagenesis screen, we isolated a new zebrafish mutant, fantome (fan), which displays a number of developmental defects including almost complete absence of cranial cartilages. A premature stop codon mutation at amino acid 356 in zebrafish wdr43 gene, the ortholog to yeast Utp5 known to function in ribosome biogenesis, is responsible for the fan phenotype. fan mutants exhibit increased apoptosis, which can be ameliorated by targeted depletion of p53 resulting in partial rescue of the craniofacial phenotype. In vitro and in vivo studies were used to demonstrate that Wdr43 is required for the proper subnucleolar localization of Tcof1, the gene often mutated in Treacher Collins syndrome, a rare autosomal dominant congenital disorder characterized by severe craniofacial abnormalities. We anticipate that the zebrafish fan mutant will be a useful tool for devising effective methods to prevent and/or treat a variety of craniofacial ribosomopathy mutations, including Treacher Collins Syndrome. This research was supported by NIH/NIDCR R01DE018043 (PCY), NIH/NIGMS R01GM52581 (SJB) and Tufts University School of Dental Medicine, Boston, MA. Astyanax mexicanus—A novel model of tooth shape formation and regeneration Despite advances in the knowledge of tooth morphogenesis and differentiation, relatively little is known about tooth regeneration which is becoming more important with the aging human population. The popular mouse model has an evolutionarily derived dentition that includes continuously growing incisors and permanent molars. The teleost fish are becoming more useful in the studies of tooth replacement and regeneration as they have teeth which are being continuously replaced throughout life. We study a relatively model animal in evolutionary development Mexican tetra (Astyanax mexicanus), which has teeth on various locations such as oral jaws, pharyngeal bones and on the gill rakers. The adult oral dentition has a unicuspid to multicuspid transition, two rows of teeth in the pre maxillary bone and late tooth formation of the maxillary bone which make this animal a good model to study the complex processes of vertebrate odontogenesis. Using this model, we investigate the molecular mechanisms behind the early stages of tooth formation, morphological transition of primary dentition to adult dentition, and tooth replacement. Whole mount bone staining, histology and in situ hybridization was done to identify the different stages of dentition development in the oral jaws. The major signaling molecules such as Shh, Bmps, and Fgf8 are well conserved in the early stages of tooth development, during tooth shape changes and at the tooth replacement stages. Future findings of this study would provide insight into biomedical research pertaining to cuspal transitions and tooth replacement cycles. This study was funded by the Natural Sciences and Engineering Research Council (NSERC), Canada. Role of Ellis-van Creveld syndrome2 (Evc2) in craniofacial development Ellis-van Creveld (EvC) syndrome is a rare chondro-ectodermal dysplasia with an autosomal recessive trait affecting bone and cartilage growth. EvC patients have mutations in either EVC or EVC2 gene, both of which are located on chromosome 4 in a head-to-head configuration. There were several cases reported that abnormal craniofacial bone phenotype was observed in EvC patients; however, it is currently unknown whether mutation of EVC or EVC2 gene causes such craniofacial bone phenotypes. In this study we used Evc2 knockout (KO) mice as an animal model of this disease and craniofacial bone development/phenotype in these mice was investigated in comparison with that of controls, that is, wild type and heterozygous mice. Our results showed that the postnatal bone growth deficiency in KO mice was found in the areas where the expression of Evc2 was observed. Growth rate of craniofacial bones in KO mice was reduced to 72–79% of that of the controls at the tested time points. Notably, growth of certain bones including nasal bone, palatal bone and premaxilla was more affected in KO than in the controls. Furthermore, there was a remarkable change in facial bones' spatial relationship to the cranial base and vault. We also found an earlier onset of apoptosis and proliferation defects in chondrocytes in KO compared to the controls. In conclusion, Evc2 is required for craniofacial bone development and deficiency in Evc2 leads to specific facial bone growth defect due to imbalance of cellular proliferation and cell death. Supported by NIH/NIDCR RO1DE019527 grant. Co-evolutionary patterning of teeth and taste buds Teeth and taste buds are iteratively patterned structures that line the oro-pharynx of many vertebrates, suggesting these structures may share both evolutionary and genetic origins. Biologists do not understand how they develop distinctly from common epithelium, or how variation in density of these units is generated. We have identified a positive correlation between spatial densities of oral teeth and taste papillae in Lake Malawi cichlid fishes. We mapped the genetic basis of density variance in an intercross between a planktivore (few teeth, few taste papillae) and algivore (many teeth, many taste papillae). Our experimental design (350 informative Rad-Tag SNPs and 300 F2) has uncovered 8 QTL that map to tooth density and 5 that map to taste papillae density. Of the ∼400 genes within 1MB of either side of these QTL, we have filtered 21 candidates of interest. These candidates are known to interact with BMP, Hh, and/or Wnt pathways which orchestrate oral organ patterning in other organisms. We assayed expression of the most intriguing candidates, along with members of the aforementioned pathways in both organs across ontogeny. Finally we delivered in vivo small molecules that perturb these pathways to affect oral organ density, some treatments of which phenocopy the natural diversity present in the Lake Malawi flock. Changes in gene expression following treatment indicate that our candidates may act in the co-patterning of teeth and taste papillae. Analysis of susceptibility loci for nonsyndromic orofacial clefting in a European trio sample This abstract is currently in press in the American Journal of Medical Genetics. Lessons from toothless jaws about dental-gnathic morphogenesis Teeth and jaws are closely linked together by position and function yet the putative influence of the developing dentition on the growing jaw skeleton is not well characterized. The p63 mouse mutant is an ideal model to probe the level of developmental interdependence between the teeth and the jaw because their morphogenesis are normal in p63+/+ and p63−/+ mice while odontogenesis, but not jaw morphogenesis, fails in p63−/− mice. As this null mutation is lethal, birth is the latest stage at which the developmental morphology of p63−/− mice can be assessed. Building on our recent study of mandibular morphology in perinatal (E18) and adult p63+/+, p63−/+ and p63−/− mice, here we investigate the effect of the loss of one and both p63 alleles on upper jaw skeletal morphology in adult (>30 days) and neonatal (P0) mice. Using a micro-computed tomography scanner (Skyscan 1172) and AMIRA software (VSG3D), we visualized and landmarked in 3D the ventral cranium/upper jaw skeleton of adult p63+/+ (n = 48) and p63−/+ (n = 60) mice, as well as neonatal p63+/+ (n = 7), p63−/+ (n = 13) and p63−/− (n = 4) mice. Following Generalized Procrustes registration of the landmark data, Principal Component analyses (morphologika 2.0) showed that, similar to the lower jaw, a single p63 allele was sufficient for the adult upper jaw to form normally. Pilot analyses of the neonatal upper jaws suggest that morphology is normal in p63+/+ and p63−/+ mice but not in p63−/− mice (P0). Our study suggests that within a p63 regulatory network, jaw morphogenesis is independent of odontogenesis. Supported by NSERC Discovery Grant #402148; and the College of Medicine, University of Saskatchewan. Cellular, molecular and genetic basis for the craniofacial phenotype of the avian mutant, talpid2 The chicken talpid2 is an autosomal recessive mutant with a myriad of congenital malformations, including facial clefting. Although phenotypically similar to its sister strain talpid3, talpid2 has a distinct facial phenotype and an unknown molecular and genetic basis. We set out to characterize the talpid2 craniofacial phenotype and determine the cellular, molecular and genetic etiology of the mutant. Cellularly, we detected aberrant levels of apoptosis localized within the maxillary mesenchyme. Molecularly, we found disruptions to the Hedgehog pathway indicative of disrupted primary cilia. Post-translational processing of Gli2 and Gli3 was aberrant in the developing facial prominences. We found that whereas both Gli2 and Gli3 processing were disrupted in talpid2 mutants, only nuclear Gli3A levels were significantly altered between control and talpid2 embryos. Finally, we identified the talpid2 phenotype as being linked to a 1.4 Mb loci on GGA1q that contains the ciliary protein C2CD3. These results suggest a cellular, molecular and genetic cause for the talpid2 craniofacial phenotype and surmise that the talpid2, whereas similar in molecular etiology to talpid3, is caused by a separate and distinct mutation in a ciliary protein. This work is supported by NIH/NIDCR 5R00DE019853. Novel IRF6 mutations in families with Van Der Woude Syndrome and Popliteal Pterygium Syndrome from Sub-Saharan Africa Orofacial clefts (OFC) are complex genetic traits that are often classified as syndromic or non-syndromic clefts. To date, over 500 syndromic clefts have been identified and included in the Online Mendelian in Man (OMIM) database and for many of these; the underlying genetic factors have been identified. Van der Woude syndrome (VWS) is a dominant disorder affecting 1/40,000 people worldwide and also one of the most common syndromic cleft, accounting for 2% of all OFC. Popliteal pterygium syndrome (PPS) is a rare dominant disorder affecting 1/300,000 people worldwide and thought to be a severe form of VWS. Mutations in the IRF6 gene have been reported to cause VWS and PPS world-wide. We obtained saliva samples from four families with VWS and one family with PPS from Nigeria and Ethiopia for Sanger sequencing of the high-risk exons for IRF6 (exons 3, 4, 7, and 9. For the VWS families, we found a novel nonsense mutation in exon 4 (p.Lys66X), a novel splice site mutation in exon 4 (p.Pro126Pro) and a previously reported splice mutation in exon 7 that changes the acceptor splice site. A previously known missense mutation was found in exon 4 (p.Arg84His) in the PPS family. All the mutations segregate in the families. The presence of IRF6 mutations in these families confirms the geographical spread of the diseases and similarity in the type of mutations. This is important for counseling and prenatal diagnosis for high-risk families especially in Africa where there are limited genetic counselors. This study was support by the NIH/NIDCR Grants K99/R00 5K99-DE022378-02, R37-DE08559, and Face Base grant U01 DE-20057. Obliteration of the intersphenoid synchondrosis affects cranial base angle but not cranial base and midface outgrowth in the sbse mouse mutant Cranial base synchondroses are thought to be significant growth sites, and premature fusion is thought to affect the growth of both the neurocranium and viscerocranium. Here, we tested the hypothesis that premature obliteration of the intersphenoid synchondrosis, the most anterior of the two midline cranial base synchondroses, contributes significantly to midface outgrowth. sbse (small body, short ear pinna) is a mutant mouse strain that exhibits a variable phenotypic spectrum characterized by significant midface hypoplasia, snout deviation, premature facial suture fusion, as well as associated doming of the cranial vault. Homozygotes are distinguishable from heterozygote and wildtype C57BL/6J littermates at an early age by their smaller size, lower birth weight and the invariable fusion of the intersphenoid synchondrosis, which begins around postnatal day (PN) 7 [cf. >PN28 in controls]. Standard linear distances of the cranial base and midface were m