Ciliopathic micrognathia is caused by aberrant skeletal differentiation and remodeling

AbstractCiliopathies represent a growing class of diseases caused by defects in microtubule-based organelles called primary cilia. Approximately 30% of ciliopathies can be characterized by craniofacial phenotypes such as craniosynostosis, cleft lip/palate and micrognathia. Patients with ciliopathic micrognathia experience a particular set of difficulties including impaired feeding and breathing and have extremely limited treatment options. To understand the cellular and molecular basis for ciliopathic micrognathia, we utilized the talpid2 (ta2), a bona fide avian model for the human ciliopathy Oral-Facial-Digital syndrome subtype 14 (OFD14). Histological analyses revealed that the onset of ciliopathic micrognathia in ta2 embryos occurred at the earliest stages of mandibular development. Neural crest-derived skeletal progenitor cells were particularly sensitive to a ciliopathic insult, undergoing unchecked passage through the cell cycle and subsequent increased proliferation. Furthermore, whereas neural crest-derived skeletal differentiation was initiated, osteoblast maturation failed to progress to completion. Additional molecular analyses revealed that an imbalance in the ratio of bone deposition and resorption also contributed to ciliopathic micrognathia in ta2 embryos. Thus, our results suggest that ciliopathic micrognathia is a consequence of multiple, aberrant cellular processes necessary for skeletal development, and provide potential avenues for future therapeutic treatments.

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Ciliopathic micrognathia is caused by aberrant skeletal differentiation and remodeling

Development ◽

10.1242/dev.194175 ◽

2021 ◽

Vol 148 (4) ◽

pp. dev194175

Author(s):

Christian Louis Bonatto Paese ◽

Evan C. Brooks ◽

Megan Aarnio-Peterson ◽

Samantha A. Brugmann

Keyword(s):

Neural Crest ◽

Cleft Lip ◽

Primary Cilia ◽

Skeletal Development ◽

Treatment Options ◽

Cellular Processes ◽

Cleft Lip Palate ◽

Bona Fide ◽

Bone Deposition ◽

Osteoblast Maturation

ABSTRACTCiliopathies represent a growing class of diseases caused by defects in microtubule-based organelles called primary cilia. Approximately 30% of ciliopathies are characterized by craniofacial phenotypes such as craniosynostosis, cleft lip/palate and micrognathia. Patients with ciliopathic micrognathia experience a particular set of difficulties, including impaired feeding and breathing, and have extremely limited treatment options. To understand the cellular and molecular basis for ciliopathic micrognathia, we used the talpid2 (ta2), a bona fide avian model for the human ciliopathy oral-facial-digital syndrome subtype 14. Histological analyses revealed that the onset of ciliopathic micrognathia in ta2 embryos occurred at the earliest stages of mandibular development. Neural crest-derived skeletal progenitor cells were particularly sensitive to a ciliopathic insult, undergoing unchecked passage through the cell cycle and subsequent increased proliferation. Furthermore, whereas neural crest-derived skeletal differentiation was initiated, osteoblast maturation failed to progress to completion. Additional molecular analyses revealed that an imbalance in the ratio of bone deposition and resorption also contributed to ciliopathic micrognathia in ta2 embryos. Thus, our results suggest that ciliopathic micrognathia is a consequence of multiple aberrant cellular processes necessary for skeletal development, and provide potential avenues for future therapeutic treatments.

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Glycosylphosphatidylinositol Biosynthesis and Remodeling are Required for Neural Crest Cell, Cardiac and Neural Development

10.1101/513507 ◽

2019 ◽

Author(s):

Marshall Lukacs ◽

Tia Roberts ◽

Praneet Chatuverdi ◽

Rolf W. Stottmann

Keyword(s):

Neural Crest ◽

Neural Development ◽

Cleft Lip ◽

Cleft Lip And Palate ◽

Folate Receptor ◽

Neural Crest Cell ◽

Specific Expression ◽

Hypomorphic Mutation ◽

Pathogenic Variants ◽

Cleft Lip Palate

AbstractThe glycosylphosphatidylinositol (GPI) anchor attaches nearly 150 proteins to the cell surface. Patients with pathogenic variants in GPI biosynthetic pathway genes display an array of phenotypes including seizures, developmental delay, dysmorphic facial features and cleft palate. There is virtually no mechanism to explain these phenotypes. we identified a novel mouse mutant (cleft lip/palate, edema and exencephaly; Clpex) with a hypomorphic mutation in Post-Glycophosphatidylinositol Attachment to Proteins-2 (Pgap2). Pgap2 is one of the final proteins in the GPI biosynthesis pathway and is required for anchor maturation. We found the Clpex mutation results in a global decrease in surface GPI expression. Surprisingly, Pgap2 showed tissue specific expression with enrichment in the affected tissues of the Clpex mutant. We found the phenotype in Clpex mutants is due to apoptosis of neural crest cells (NCCs) and the cranial neuroepithelium, as is observed in the GPI anchored Folate Receptor 1-/- mouse. We showed folinic acid supplementation in utero can rescue the cleft lip phenotype in Clpex. Finally, we generated a novel mouse model of NCC-specific total GPI deficiency in the Wnt1-Cre lineage. These mutants developed median cleft lip and palate demonstrating a cell autonomous role for GPI biosynthesis in NCC development.

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Genetics of Dentofacial and Orthodontic Abnormalities

Global Medical Genetics ◽

10.1055/s-0040-1722303 ◽

2021 ◽

Author(s):

Praveen Kumar Neela ◽

Anjana Atteeri ◽

Pavan Kumar Mamillapalli ◽

Vasu Murthy Sesham ◽

Sreekanth Keesara ◽

...

Keyword(s):

Cleft Lip ◽

Experimental Studies ◽

Twin Studies ◽

Pedigree Analysis ◽

Cellular Processes ◽

Treatment Techniques ◽

Cleft Lip Palate ◽

Genetic Mechanisms ◽

Genetic And Environmental Factors

AbstractThe development of craniofacial complex and dental structures is a complex and delicate process guided by specific genetic mechanisms. Genetic and environmental factors can influence the execution of these mechanisms and result in abnormalities. An insight into the mechanisms and genes involved in the development of orofacial and dental structures has gradually gained by pedigree analysis of families and twin studies as well as experimental studies on vertebrate models. The development of novel treatment techniques depends on in-depth knowledge of the various molecular or cellular processes and genes involved in the development of the orofacial complex. This review article focuses on the role of genes in the development of nonsyndromic orofacial, dentofacial variations, malocclusions, excluding cleft lip palate, and the advancements in the field of molecular genetics and its application to obtain better treatment outcomes.

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Cyclopamine-Induced Holoprosencephaly and Associated Craniofacial Malformations in the Golden Hamster: Anatomic and Molecular Events

Pediatric and Developmental Pathology ◽

10.1007/s100249900004 ◽

1998 ◽

Vol 1 (1) ◽

pp. 29-41 ◽

Cited By ~ 24

Author(s):

Susan Coventry ◽

Raj P. Kapur ◽

Joseph R. Siebert

Keyword(s):

Neural Crest ◽

Cleft Lip ◽

Neural Plate ◽

Early Event ◽

Desert Plant ◽

Craniofacial Malformations ◽

Altered Expression ◽

Facial Anomalies ◽

Molecular Events ◽

Cleft Lip Palate

Holoprosencephaly is a complex congenital malformation of the brain and is often associated with a spectrum of facial anomalies ranging from normocephaly or nondiagnostic changes to cleft lip/palate (premaxillary dysgenesis), cebocephaly ethmocephaly and cyclopia. The primary insult is thought to occur during gastrulation, when prechordal mesenchyme and overlying anterior neural plate undergo complex developmental interactions. Exposure to cyclopamine, a steroid isolated from the desert plant Veratrum californicum, causes holoprosencephaly in mammalian embryos. We have begun to study the pathogenesis of cyclopamine-induced holoprosencephaly and associated craniofacial anomalies in Syrian golden hamsters ( Mesocricetus auratus). Embryos were exposed to a single maternal dose of cyclopamine during gastrulation on embryonic day (E) 7.0. By E13.0, 62% of fetuses showed craniofacial malformations, including premaxillary dysgenesis, ocular hypotelorism, and cebocephaly. Facial anomalies were associated with absence of the premaxilla and abnormalities of the midline cranial base, particularly the ethmoid and sphenoid bones. Histological sections from cyclopamine-treated embryos at earlier stages showed marked deficiency of cranial mesenchyme derived from the rostral neural crest. Expression of two transcription factors, HNF-3β and Hox-b5, which have been implicated in specification of rostral and caudal neural crest cells, respectively, were examined immunohistochemically. Treatment with cyclopamine caused a transient loss of HNF-3β immunoreactivity in prechordal mesenchyme, but had no effect on Hox-b5 expression. The findings suggest that an early event in the pathogenesis of cyclopamine-induced holoprosencephaly may be altered expression of selected proteins in the prechordal mesenchyme and floor plate with secondary impaired development of the adjacent neural plate and cranial neural crest.

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Disrupted IRF6-NME1/2 Complexes as a Cause of Cleft Lip/Palate

Journal of Dental Research ◽

10.1177/0022034517723615 ◽

2017 ◽

Vol 96 (11) ◽

pp. 1330-1338 ◽

Cited By ~ 8

Author(s):

M.T. Parada-Sanchez ◽

E.Y. Chu ◽

L.L. Cox ◽

S.S. Undurty ◽

J.M. Standley ◽

...

Keyword(s):

Epithelial Cells ◽

Cleft Lip ◽

Missense Mutations ◽

C Terminus ◽

Interferon Regulatory Factor 6 ◽

Cleft Lip Palate ◽

Bona Fide ◽

Partner Proteins

Mutations and common polymorphisms in interferon regulatory factor 6 ( IRF6) are associated with both syndromic and nonsyndromic forms of cleft lip/palate (CLP). To date, much of the focus on this transcription factor has been on identifying its direct targets and the gene regulatory network in which it operates. Notably, however, IRF6 is found predominantly in the cytoplasm, with its import into the nucleus tightly regulated like other members of the IRF family. To provide further insight into the role of IRF6 in the pathogenesis of CLP, we sought to identify direct IRF6 protein interactors using a combination of yeast 2-hybrid screens and co-immunoprecipitation assays. Using this approach, we identified NME1 and NME2, well-known regulators of Rho-type GTPases, E-cadherin endocytosis, and epithelial junctional remodeling, as bona fide IRF6 partner proteins. The NME proteins co-localize with IRF6 in the cytoplasm of primary palatal epithelial cells in vivo, and their interaction with IRF6 is significantly enhanced by phosphorylation of key serine residues in the IRF6 C-terminus. Furthermore, CLP associated IRF6 missense mutations disrupt the ability of IRF6 to bind the NME proteins and result in elevated activation of Rac1 and RhoA, compared to wild-type IRF6, when ectopically expressed in 293T epithelial cells. Significantly, we also report the identification of 2 unique missense mutations in the NME proteins in patients with CLP (NME1 R18Q in an IRF6 and GRHL3 mutation-negative patient with van der Woude syndrome and NME2 G71V in a patient with nonsyndromic CLP). Both variants disrupted the ability of the respective proteins to interact with IRF6. The data presented suggest an important role for cytoplasmic IRF6 in regulating the availability or localization of the NME1/2 complex and thus the dynamic behavior of epithelia during lip/palate development.

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Canonical and noncanonical intraflagellar transport regulates craniofacial skeletal development

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1519458113 ◽

2016 ◽

Vol 113 (19) ◽

pp. E2589-E2597 ◽

Cited By ~ 33

Author(s):

Kazuo Noda ◽

Megumi Kitami ◽

Kohei Kitami ◽

Masaru Kaku ◽

Yoshihiro Komatsu

Keyword(s):

Neural Crest ◽

Intracellular Transport ◽

Primary Cilia ◽

Type I Collagen ◽

Skeletal Development ◽

Growth Factor Receptor ◽

Intraflagellar Transport ◽

Type I ◽

Multipotent Stem Cells ◽

Facial Region

The primary cilium is a cellular organelle that coordinates signaling pathways critical for cell proliferation, differentiation, survival, and homeostasis. Intraflagellar transport (IFT) plays a pivotal role in assembling primary cilia. Disruption and/or dysfunction of IFT components can cause multiple diseases, including skeletal dysplasia. However, the mechanism by which IFT regulates skeletogenesis remains elusive. Here, we show that a neural crest-specific deletion of intraflagellar transport 20 (Ift20) in mice compromises ciliogenesis and intracellular transport of collagen, which leads to osteopenia in the facial region. Whereas platelet-derived growth factor receptor alpha (PDGFRα) was present on the surface of primary cilia in wild-type osteoblasts, disruption ofIft20down-regulated PDGFRα production, which caused suppression of PDGF-Akt signaling, resulting in decreased osteogenic proliferation and increased cell death. Although osteogenic differentiation in cranial neural crest (CNC)-derived cells occurred normally inIft20-mutant cells, the process of mineralization was severely attenuated due to delayed secretion of type I collagen. In control osteoblasts, procollagen was easily transported from the endoplasmic reticulum (ER) to the Golgi apparatus. By contrast, despite having similar levels of collagen type 1 alpha 1 (Col1a1) expression,Ift20mutants did not secrete procollagen because of dysfunctional ER-to-Golgi trafficking. These data suggest that in the multipotent stem cells of CNCs, IFT20 is indispensable for regulating not only ciliogenesis but also collagen intracellular trafficking. Our study introduces a unique perspective on the canonical and noncanonical functions of IFT20 in craniofacial skeletal development.

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Glycosylphosphatidylinositol biosynthesis and remodeling are required for neural tube closure, heart development, and cranial neural crest cell survival

eLife ◽

10.7554/elife.45248 ◽

2019 ◽

Vol 8 ◽

Cited By ~ 7

Author(s):

Marshall Lukacs ◽

Tia Roberts ◽

Praneet Chatuverdi ◽

Rolf W Stottmann

Keyword(s):

Neural Crest ◽

Heart Development ◽

Cleft Lip ◽

Cleft Lip And Palate ◽

Neural Crest Cell ◽

Neural Tube Closure ◽

Specific Expression ◽

Pathogenic Variants ◽

Median Cleft ◽

Cleft Lip Palate

Glycosylphosphatidylinositol (GPI) anchors attach nearly 150 proteins to the cell membrane. Patients with pathogenic variants in GPI biosynthesis genes develop diverse phenotypes including seizures, dysmorphic facial features and cleft palate through an unknown mechanism. We identified a novel mouse mutant (cleft lip/palate, edema and exencephaly; Clpex) with a hypo-morphic mutation in Post-Glycophosphatidylinositol Attachment to Proteins-2 (Pgap2), a component of the GPI biosynthesis pathway. The Clpex mutation decreases surface GPI expression. Surprisingly, Pgap2 showed tissue-specific expression with enrichment in the brain and face. We found the Clpex phenotype is due to apoptosis of neural crest cells (NCCs) and the cranial neuroepithelium. We showed folinic acid supplementation in utero can partially rescue the cleft lip phenotype. Finally, we generated a novel mouse model of NCC-specific total GPI deficiency. These mutants developed median cleft lip and palate demonstrating a previously undocumented cell autonomous role for GPI biosynthesis in NCC development.

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