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.

National Institute of Health (NIH)
National Institute of Dental & Craniofacial Research (NIDCR)
Career Transition Award (K22)
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NIDCR Special Grants Review Committee (DSR)
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Hardwick, Kevin S
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Forsyth Institute
United States
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Spencer Adams, Dany; Lemire, Joan M; Kramer, Richard H et al. (2014) Optogenetics in Developmental Biology: using light to control ion flux-dependent signals in Xenopus embryos. Int J Dev Biol 58:851-61
Adams, Dany S; Levin, Michael (2013) Endogenous voltage gradients as mediators of cell-cell communication: strategies for investigating bioelectrical signals during pattern formation. Cell Tissue Res 352:95-122
Adams, Dany S; Levin, Michael (2012) General principles for measuring resting membrane potential and ion concentration using fluorescent bioelectricity reporters. Cold Spring Harb Protoc 2012:385-97
Vandenberg, Laura N; Adams, Dany S; Levin, Michael (2012) Normalized shape and location of perturbed craniofacial structures in the Xenopus tadpole reveal an innate ability to achieve correct morphology. Dev Dyn 241:863-78
Chernet, Brook T; Adams, Dany S; Levin, Michael (2012) Photoconversion for tracking the dynamics of cell movement in Xenopus laevis embryos. Cold Spring Harb Protoc 2012:683-90
Adams, Dany S; Levin, Michael (2012) Measuring resting membrane potential using the fluorescent voltage reporters DiBAC4(3) and CC2-DMPE. Cold Spring Harb Protoc 2012:459-64
Mondia, Jessica P; Levin, Michael; Omenetto, Fiorenzo G et al. (2011) Long-distance signals are required for morphogenesis of the regenerating Xenopus tadpole tail, as shown by femtosecond-laser ablation. PLoS One 6:e24953
Blackiston, Douglas; Adams, Dany S; Lemire, Joan M et al. (2011) Transmembrane potential of GlyCl-expressing instructor cells induces a neoplastic-like conversion of melanocytes via a serotonergic pathway. Dis Model Mech 4:67-85
Beane, Wendy S; Morokuma, Junji; Adams, Dany S et al. (2011) A chemical genetics approach reveals H,K-ATPase-mediated membrane voltage is required for planarian head regeneration. Chem Biol 18:77-89
Vandenberg, Laura N; Morrie, Ryan D; Adams, Dany Spencer (2011) V-ATPase-dependent ectodermal voltage and pH regionalization are required for craniofacial morphogenesis. Dev Dyn 240:1889-904

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