Department of Cell and Developmental Biology, University of North Carolina at Chapel Hill School of Medicine, Chapel Hill, North Carolina, United States of America.
PLoS One. 2011;6(6):e20309. doi: 10.1371/journal.pone.0020309. Epub 2011 Jun 8.
Yes-associated protein 65 (YAP) contains multiple protein-protein interaction domains and functions as both a transcriptional co-activator and as a scaffolding protein. Mouse embryos lacking YAP did not survive past embryonic day 8.5 and showed signs of defective yolk sac vasculogenesis, chorioallantoic fusion, and anterior-posterior (A-P) axis elongation. Given that the YAP knockout mouse defects might be due in part to nutritional deficiencies, we sought to better characterize a role for YAP during early development using embryos that develop externally. YAP morpholino (MO)-mediated loss-of-function in both frog and fish resulted in incomplete epiboly at gastrulation and impaired axis formation, similar to the mouse phenotype. In frog, germ layer specific genes were expressed, but they were temporally delayed. YAP MO-mediated partial knockdown in frog allowed a shortened axis to form. YAP gain-of-function in Xenopus expanded the progenitor populations in the neural plate (sox2(+)) and neural plate border zone (pax3(+)), while inhibiting the expression of later markers of tissues derived from the neural plate border zone (neural crest, pre-placodal ectoderm, hatching gland), as well as epidermis and somitic muscle. YAP directly regulates pax3 expression via association with TEAD1 (N-TEF) at a highly conserved, previously undescribed, TEAD-binding site within the 5' regulatory region of pax3. Structure/function analyses revealed that the PDZ-binding motif of YAP contributes to the inhibition of epidermal and somitic muscle differentiation, but a complete, intact YAP protein is required for expansion of the neural plate and neural plate border zone progenitor pools. These results provide a thorough analysis of YAP mediated gene expression changes in loss- and gain-of-function experiments. Furthermore, this is the first report to use YAP structure-function analyzes to determine which portion of YAP is involved in specific gene expression changes and the first to show direct in vivo evidence of YAP's role in regulating pax3 neural crest expression.
Yes 相关蛋白 65（YAP）包含多个蛋白-蛋白相互作用结构域，作为转录共激活因子和支架蛋白发挥作用。缺乏 YAP 的小鼠胚胎在胚胎第 8.5 天之前不能存活，并表现出卵黄囊血管生成、绒毛膜尿囊融合和前后（A-P）轴伸长缺陷的迹象。由于 YAP 敲除小鼠的缺陷可能部分是由于营养缺乏所致，我们试图使用外部发育的胚胎更好地描述 YAP 在早期发育中的作用。在青蛙和鱼类中，YAP 形态发生素（MO）介导的功能丧失导致原肠胚形成时不完全内卷和轴形成受损，类似于小鼠表型。在青蛙中，胚层特异性基因被表达，但它们的时间延迟。在青蛙中，YAP MO 介导的部分敲低允许形成缩短的轴。Xenopus 中的 YAP 功能获得扩大了神经板（sox2(+))和神经板边界区（pax3(+))中的祖细胞群体，同时抑制了神经板边界区衍生组织的后期标记物（神经嵴、前脑嵴外胚层、孵化腺）以及表皮和体节肌肉的表达。YAP 通过与 TEAD1（N-TEF）在 pax3 的 5'调控区的一个高度保守的、以前未描述的 TEAD 结合位点结合，直接调节 pax3 的表达。结构/功能分析表明，YAP 的 PDZ 结合基序有助于抑制表皮和体节肌肉分化，但完整的、完整的 YAP 蛋白是扩展神经板和神经板边界区祖细胞池所必需的。这些结果提供了在缺失和获得功能实验中对 YAP 介导的基因表达变化的全面分析。此外，这是首次使用 YAP 结构-功能分析来确定 YAP 的哪一部分参与特定基因表达变化的报告，也是首次显示 YAP 在调节 pax3 神经嵴表达中的直接体内证据。
