Dyrk1a dysregulation in Trisomy 21 and Dyrk1a haploinsufficiency lead to midface hypoplasia

21号染色体三体是由21号染色体的部分或完全重复引起的。这种疾病导致智力障碍， 残疾、心脏缺陷和明显的颅面畸形。一些研究调查了 这些缺陷的遗传来源。一个候选基因是双特异性激酶Dyrk1a。Dyrk1a是一个有吸引力的 Dyrk1a单倍不足也会导致颅面缺损。而颅面 由这些遗传不平衡引起的缺陷是不同的，一个共同的特征是面中部发育不全。先前 我们实验室的工作已经确定非洲爪蟾是研究面中部发育的理想生物 和初级腭。我们的实验室和其他实验室先前的研究表明，dyrk1a表达于 非洲爪蟾的脸我假设Dyrk1a是面中部发育的重要调节因子， dyrk1a的剂量不平衡导致21三体和Dryk1a中存在的不同特征 单倍不足为了验证这一假设，我将利用非洲爪蟾检查dyrk1a的表达， 口面发育我将研究dyrk1a减少的影响，通过药理学抑制， INDY，Dyrk1a特异性抑制剂，或反义寡核苷酸吗啉（Dyrk1a MO）。评估 过表达的影响我将注射一个经过验证的全长爪蟾dyrk1a mRNA构建体。我必坚定 简单的面部测量和几何形态测量，以定性和定量地研究 dyrk1a剂量不平衡的影响。Dyrk1a信号在面部的细胞需求和机制 仍有待阐明。我将通过仔细操作INDY治疗窗口来探索这一点， 神经嵴规格、迁移、分层和分化的评估。我希望找到一个角色 在一个或多个这些过程中的Dyrk1a。为了进一步研究Dyrk1a在面部的要求，我 将用dyrk1a MO进行面部移植试验。在目标2中，我将研究视黄酸的作用 (RA)在Dyrk1a的调节中。Dyrk1a被我们的实验室确定为RA的靶点，在一个大的基因组筛选中， 发育中的胚胎RA信号减弱。RA的剂量失衡，无论是增加还是 减少，导致面中部发育不全和面中部裂。我假设RA抑制了 dyrk1a的表达。已知RA介导的抑制调节发育中胚胎中fgf 8的水平， 我们实验室的初步数据表明，RA的抑制可以与dyrk1a协同作用， 异常我将在过量RA范例中检查dryk1a水平，并检查是否人为调节RA 水平诱导dyrk1a的预期变化。如果是这种情况，RA增加将导致dyrk1a降低 且反之亦然。我将通过研究强迫剂量失衡的可能性来结束这一目标， 增加RA的剂量，挽救dyrk1a增加诱导的面中部表型。这些 研究将进一步了解Dyrk1a在面中部发育过程中的作用， 与21三体和Dyrk1a单倍不足相关的面中部异常。

Trisomy 21 results from partial or complete duplication of chromosome 21. This disorder results in intellectual disabilities, cardiac defects, and distinct craniofacial abnormalities. Several studies have investigated the genetic origin of these defects. A candidate gene is the dual specificity kinase, Dyrk1a. Dyrk1a is an attractive target, given that Dyrk1a haploinsufficiency results in craniofacial defects as well. While the craniofacial defects induced by these genetic imbalances are distinct, a common feature is midface hypoplasia. Previous work in our laboratory has established Xenopus laevis as ideal organism to study development of the midface and primary palate. Previous investigations, by our laboratory and others, suggest that dyrk1a is expressed in the Xenopus face. I hypothesize that Dyrk1a is an important regulator of midface development and a dosage imbalance of dyrk1a results in the distinct features present in Trisomy 21 and Dryk1a haploinsufficiency. To test this hypothesis, I will utilize Xenopus to examine the expression of dyrk1a during orofacial development. I will investigate the effects of decreased dyrk1a, via pharmacological inhibition with INDY, a Dyrk1a-specific inhibitor, or antisense oligonucleotide morpholinos (Dyrk1a MO). To assess the effects of overexpression I will inject a verified, full-length Xenopus dyrk1a mRNA construct. I will perform simple face measurements and geometric morphometrics to qualitatively and quantitatively investigate the effects of dyrk1a dosage imbalance. The cellular requirements and mechanisms of Dyrk1a signaling in the face remain to be elucidated. I will explore this by careful manipulation of the INDY treatment window and assessment of neural crest specification, migration, delamination, and differentiation. I expect to identify a role of Dyrk1a in one or more of these processes. To further investigate the requirements of Dyrk1a in the face, I will perform face transplantation assays with the dyrk1a MO. In Aim 2, I will investigate the role of retinoic acid (RA) in the regulation of Dyrk1a. Dyrk1a was identified our lab as a target of RA, in a large genomic screen of developing embryos with diminished RA signaling. A dosage imbalance of RA, whether increased or decreased, results in midfacial hypoplasia and a midfacial cleft. I hypothesize that RA inhibits the expression of dyrk1a. RA-mediated repression is known regulate levels of fgf8 in the developing embryo and preliminary data from our lab suggest that inhibition of RA can synergize with dyrk1a to produce facial abnormalities. I will examine dryk1a levels in an excess RA paradigm, and examine if artificially modulating RA levels induces the expected change in dyrk1a. If this is the case, increased RA will result in decreased dyrk1a and vice versa. I will conclude this aim by investigating the possibility of forcing the dosage imbalance, increasing the dosage of RA, and rescuing the midfacial phenotype induced by increased dyrk1a. These studies will further the understanding of the role of Dyrk1a during midfacial development and provide insight into the midfacial abnormalities associated with Trisomy 21 and Dyrk1a haploinsufficiency.