Ductin's Role in Craniofacial Patterning and Development

描述（由申请人提供）：最近的实验提供了强有力的证据，证明蛋白质ductin在颅面发育中起重要作用。Ductin也被称为v - atp酶的16kD亚基c，其功能是将质子从细胞质中跨膜运输到囊泡或细胞外空间；Ductin也可能单独或与连接蛋白一起形成间隙连接。因此，导管蛋白在细胞内、细胞间和细胞外的离子通量中起着重要的作用。我们最近发现，在非洲爪蟾胚胎中错误表达野生型导管蛋白会导致颅面异常，表达显性阴性导管蛋白或注射抗导管蛋白抗体也是如此。处理过的胚胎的表型，包括畸形的眼睛、耳朵和头骨，看起来与注射抗连接蛋白抗体和错误表达爪蟾卷曲-3蛋白引起的胚胎非常相似。我们已经知道ductin在Sonic hedgehog信号通路的上游起作用；我们的新结果提出了一个令人着迷的前景，即在脊椎动物头部形态发生过程中，传导蛋白和离子通量也在wnt卷曲级联的上游。我们建议探讨假设

DESCRIPTION (provided by applicant): Recent experiments provide strong evidence that the protein ductin plays an important role in craniofacial development. Ductin, also known as the 16kD subunit c of V-ATPase, functions in transmembrane transport of protons out of the cytoplasm and into vesicles or the extracellular space; ductin can probably also form gap junctions either alone or with connexins. Thus, ductin has important roles in ion flux across intra-, inter-, and extra-cellular boundaries. We recently discovered that misexpressing wildtype ductin in Xenopus embryos leads to craniofacial abnormalities, as does expressing a dominant negative ductin or injecting antiductin antibodies. The phenotypes of treated embryos, including malformed eyes, ears, and skull, look remarkably like those caused by injection of anti-connexin antibodies and misexpression of the Xenopus frizzled-3 protein. We already know that ductin functions upstream of the Sonic hedgehog signaling pathway; our new results raise the fascinating prospect that ductin and ion flux are also upstream of the Wnt-frizzled cascade during morphogenesis of the vertebrate head. We propose to explore the hypothesis that ductin-based ion flux is an early step in craniofacial morphogenesis that regulates signaling cascades via both V-ATPase dependent proton pumping and cell-cell gap junctional communication. Imaging using in situ hybridization, immuncytochemistry, and in vivo monitoring of pH and fluorescently labeled proteins will be used to characterize ductin's spatial and temporal expression pattern and to monitor the effects of ductin inhibition and misexpression. Loss and gain of function experiments using wildtype and dominant negative ductin constructs will allow us to study the relationship of ductin to downstream genetic pathways. This work will yield important and novel information about the under-studied phenomenon of biophysical control of developmental events, e.g., ion control of secreted signals, thus providing unique and valuable insight into normal morphogenesis and suggesting novel approaches to treating and preventing birth defects.