Cardiac outflow tract septation failure in Pax3-deficient embryos is due to p53-dependent regulation of migrating cardiac neural crest

Section on Developmental and Stem Cell Biology, Joslin Diabetes Center, One Joslin Place, Boston, MA 02215, USA.
Mechanisms of development (Impact Factor: 2.44). 09/2008; 125(9-10):757-67. DOI: 10.1016/j.mod.2008.07.003
Source: PubMed


During neural tube closure, Pax3 is required to inhibit p53-dependent apoptosis. Pax3 is also required for migration of cardiac neural crest (CNC) from the neural tube to the heart and septation of the primitive single cardiac outflow tract into the aorta and pulmonary arteries. Whether Pax3 is required for CNC migration and outflow tract septation by inhibiting p53-dependent apoptosis is not known. In this study, mouse strains carrying reporters linked to Pax3 alleles were used to map the fate of CNC cells in embryos which were either Pax3-sufficient (expressing one or two functional Pax3 alleles) or Pax3-deficient (expressing two null Pax3 alleles), and in which p53 had been inactivated or not. Migrating CNC cells were observed in both Pax3-sufficient and -deficient embryos, but CNC cells were sparse and disorganized in Pax3-deficient embryos as migration progressed. The defective migration was associated with increased cell death. Suppression of p53, either by null mutation of the p53 gene, or administration of a p53 inhibitor, pifithrin-alpha, prevented the defective CNC migration and apoptosis in Pax3-deficient embryos, and also restored proper development of cardiac outflow tracts. These results indicate that Pax3 is required for cardiac outflow tract septation because it blocks p53-dependent processes during CNC migration.

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Available from: Mary R Loeken, Oct 08, 2015
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    • "Via the Wnt/Fz/Dvl pathway, DAAM1 with RhoA participates in morphology and migratory behaviors in vertebrates such as cell fate specification, migration, proliferation and apoptosis [13,17]. Cardiac neural crest (cNC) passes through pharyngeal arches to the efferent pathway of the heart, and subsequent directional cell migration are essential for the formation of cardiac outflow tract [18]. Failure of the cNC directed migration may cause severe defects in the conotruncal region and the atrioventricular septum. "
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    ABSTRACT: Background With an increasing incidence of congenital heart defects (CHDs) in recent years, genotype-phenotype correlation and array-based methods have contributed to the genome-wide analysis and understanding of genetic variations in the CHD population. Here, we report a copy number deletion of chromosomal 14q23.1 in a female fetus with complex congenital heart defects. This is the first description of DAAM1 gene deletion associated with congenital heart anomalies. Case Presentation Compared with the control population, one CHD fetus showed a unique copy number deletion of 14q23.1, a region that harbored DAAM1 and KIAA0666 genes. Conclusions Results suggest that the copy number deletion on chromosome 14q23.1 may be critical for cardiogenesis. However, the exact relationship and mechanism of how DAAM1 and KIAA0666 deletion contributes to the onset of CHD is yet to be determined.
    BMC Medical Genetics 08/2012; 13(1):63. DOI:10.1186/1471-2350-13-63 · 2.08 Impact Factor
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    • "This suggests that the ensuing NTD and COTD result from depletion of progenitor cells that are necessary to populate these structures. We showed that inactivation of p53 through germ-line mutation or chemical inhibition prevented the NTD, exencephaly and spina bifida, and COTD that are characteristic of Pax3Sp/Sp embryos, as well as associated apoptosis, in embryos expressing nonfunctional Pax3 alleles [25], [27]. This indicates that Pax3 is not required in neuroepithelium and neural crest to regulate genes that direct morphogenesis or migration, but that it is required to block p53-dependent processes that lead to apoptosis. "
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    ABSTRACT: Pax3 is a developmental transcription factor that is required for neural tube and neural crest development. We previously showed that inactivating the p53 tumor suppressor protein prevents neural tube and cardiac neural crest defects in Pax3-mutant mouse embryos. This demonstrates that Pax3 regulates these processes by blocking p53 function. Here we investigated the mechanism by which Pax3 blocks p53 function. We employed murine embryonic stem cell (ESC)-derived neuronal precursors as a cell culture model of embryonic neuroepithelium or neural crest. Pax3 reduced p53 protein stability, but had no effect on p53 mRNA levels or the rate of p53 synthesis. Full length Pax3 as well as fragments that contained either the DNA-binding paired box or the homeodomain, expressed as GST or FLAG fusion proteins, physically associated with p53 and Mdm2 both in vitro and in vivo. In contrast, Splotch Pax3, which causes neural tube and neural crest defects in homozygous embryos, bound weakly, or not at all, to p53 or Mdm2. The paired domain and homeodomain each stimulated Mdm2-mediated ubiquitination of p53 and p53 degradation in the absence of the Pax3 transcription regulatory domains, whereas Splotch Pax3 did not stimulate p53 ubiquitination or degradation. Pax3 inactivates p53 function by stimulating its ubiquitination and degradation. This process utilizes the Pax3 paired domain and homeodomain but is independent of DNA-binding and transcription regulation. Because inactivating p53 is the only required Pax3 function during neural tube closure and cardiac neural crest development, and inactivating p53 does not require Pax3-dependent transcription regulation, this indicates that Pax3 is not required to function as a transcription factor during neural tube closure and cardiac neural crest development. These findings further suggest novel explanations for PAX3 functions in human diseases, such as in neural crest-derived cancers and Waardenburg syndrome types 1 and 3.
    PLoS ONE 12/2011; 6(12):e29379. DOI:10.1371/journal.pone.0029379 · 3.23 Impact Factor
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    • "Taken together, this suggests that diabetes-associated malformations of the face and heart may result from impaired NCC development caused by embryonic oxidative stress, a hypothesis we have proposed and previously scrutinized (Simán et al., 2000; Molin et al., 2004). This notion of ROS-induced NCC maldevelopment in diabetic pregnancy has found subsequent support in the literature (Roest et al., 2007; Morgan et al., 2008a,b). "
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    ABSTRACT: Diabetic pregnancy is associated with increased risk of malformation in the infant. Diabetes-induced anomalies of the face and heart are strongly correlated with neural crest cell (NCC) maldevelopment. We aimed to study glucose-induced alterations of mRNA levels in cranial and trunk NCCs isolated from rat embryos with increased risk of developing mandibular and cardiac malformations in diabetic pregnancy. Inbred Sprague-Dawley rat embryos were used for NCC isolation from neural tube explants. The migrating cells were exposed to 5.5 or 30 mmol/l glucose concentration for 48 hr, harvested, and prepared for gene expression measurement by RT-PCR or immunostaining with either distal-less (Dlx) or AP-2-α antibodies. Evaluation of the immunostained slides showed that approximately 75% of the cells were of NCC origin. Exposure to 30 mM glucose decreased mRNA levels of Copper-Zinc superoxide dismutase, manganese superoxide dismutase, extracellular superoxide dismutase, Catalase, Gpx-1, Nrf2, poly-ADP ribose polymerase, B-cell leukemia/lymphoma protein 2, and β-Catenin genes in cranial neural crest explant cultures. In addition, Pax-3, Pax-6, Wnt3a, and Apc mRNA levels were decreased by high glucose exposure in both cranial and trunk neural crest explant cultures. Cranial NCCs diminish their mRNA levels of antioxidative enzymes and the Nrf2 response factor, as well as the antiapoptotic B-cell leukemia/lymphoma protein 2 gene, in response to increased ambient glucose concentration. Furthermore, both cranial and trunk NCC decrease the mRNA levels of the transcription factors Pax-3 and Pax-6, as well as key components of the Wnt pathway. These patterns of glucose-altered gene expression in a developmentally important cell population may be of etiological importance for NCC-associated malformations in diabetic pregnancy.
    Birth Defects Research Part B Developmental and Reproductive Toxicology 10/2011; 92(5):487-97. DOI:10.1002/bdrb.20321 · 0.77 Impact Factor
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