Department of Biological Chemistry, Howard Hughes Medical Institute, University of California, Los Angeles, CA 90095–1662, USA.
Dev Biol. 2011 Apr 15;352(2):317-28. doi: 10.1016/j.ydbio.2011.01.029. Epub 2011 Feb 3.
In Xenopus, dorsal-ventral (D-V) patterning can self-regulate after embryo bisection. This is mediated by an extracellular network of proteins secreted by the dorsal and ventral centers of the gastrula. Different proteins of similar activity can be secreted at these two poles, but under opposite transcriptional control. Here we show that Crescent, a dorsal protein, can compensate for the loss of Sizzled, a ventral protein. Crescent is a secreted Frizzled-Related Protein (sFRP) known to regulate Wnt8 and Wnt11 activity. We now find that Crescent also regulates the BMP pathway. Crescent expression was increased by the BMP antagonist Chordin and repressed by BMP4, while the opposite was true for Sizzled. Crescent knock-down increased the expression of BMP target genes, and synergized with Sizzled morpholinos. Thus, Crescent loss-of-function is compensated by increased expression of its ventral counterpart Sizzled. Crescent overexpression dorsalized whole embryos but not ventral half-embryos, indicating that Crescent requires a dorsal component to exert its anti-BMP activity. Crescent protein lost its dorsalizing activity in Chordin-depleted embryos. When co-injected, Crescent and Chordin proteins greatly synergized in the dorsalization of Xenopus embryos. The molecular mechanism of these phenotypes is explained by the ability of Crescent to inhibit Tolloid metalloproteinases, which normally degrade Chordin. Enzyme kinetic studies showed that Crescent was a competitive inhibitor of Tolloid activity, which bound to Tolloid/BMP1 with a K(D) of 11 nM. In sum, Crescent is a new component of the D-V pathway, which functions as the dorsal counterpart of Sizzled, through the regulation of chordinases of the Tolloid family.
在非洲爪蟾中，背腹（D-V）模式可以在胚胎横切后自我调节。这是由原肠胚背部和腹部中心分泌的细胞外蛋白质网络介导的。具有相似活性的不同蛋白质可以在这两个极点分泌，但转录控制相反。在这里，我们表明，背侧蛋白 Crescent 可以补偿腹侧蛋白 Sizzled 的缺失。 Crescent 是一种已知调节 Wnt8 和 Wnt11 活性的分泌型 Frizzled 相关蛋白（sFRP）。我们现在发现 Crescent 也调节 BMP 途径。 Crescent 表达受 BMP 拮抗剂 Chordin 的增加和 BMP4 的抑制，而 Sizzled 的情况则相反。 Crescent 敲低增加了 BMP 靶基因的表达，并与 Sizzled 形态发生素协同作用。因此， Crescent 功能丧失通过其腹侧对应物 Sizzled 的表达增加得到补偿。 Crescent 过表达使整个胚胎背侧化，但不使腹侧半胚胎背侧化，表明 Crescent 需要背侧成分才能发挥其抗 BMP 活性。 Crescent 蛋白在 Chordin 耗尽的胚胎中失去其背侧化活性。当共同注射时，Crescent 和 Chordin 蛋白在非洲爪蟾胚胎的背侧化中大大协同作用。这些表型的分子机制可以解释为 Crescent 抑制 Tolloid 金属蛋白酶的能力，Tolloid 金属蛋白酶通常降解 Chordin。酶动力学研究表明，Crescent 是 Tolloid 活性的竞争性抑制剂，与 Tolloid/BMP1 的结合 K(D)为 11 nM。总之，Crescent 是 D-V 途径的一个新组成部分，它通过调节 Tolloid 家族的 Chordinases，作为 Sizzled 的背侧对应物发挥作用。
