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Defining pattern formation mechanisms during embryonic development is important for understanding the etiology of birth defects and to inform tissue engineering approaches. In this study, we used tricaine, a voltage-gated sodium channel (VGSC) inhibitor, to show that VGSC activity is required for normal skeletal patterning in Lytechinus variegatus sea urchin larvae. We demonstrate that tricaine-mediated patterning defects are rescued by an anesthetic-insensitive version of the VGSC LvScn5a. Expression of this channel is enriched in the ventrolateral ectoderm where it spatially overlaps with posterolaterally expressed Wnt5. We show that VGSC activity is required to spatially restrict Wnt5 expression to this ectodermal region that is adjacent and instructive to clusters of primary mesenchymal cells that initiate secretion of the larval skeleton as triradiates. Tricaine-mediated Wnt5 spatial expansion correlates with the formation of ectopic PMC clusters and triradiates. These defects are rescued by Wnt5 knock-down, indicating that the spatial expansion Wnt5 is responsible for the patterning defects induced by VGSC inhibition. These results demonstrate a novel connection between bioelectrical status and the spatial control of patterning cue expression during embryonic pattern formation. 

Abstract We provide a functional characterization of transcription factor NF-κB in protists and provide information about the evolution and diversification of this biologically important protein. We characterized NF-κB in two protists using phylogenetic, cellular, and biochemical techniques. NF-κB of the holozoanCapsaspora owczarzaki(Co) has an N-terminal DNA-binding domain and a C-terminal Ankyrin repeat (ANK) domain, and its DNA-binding specificity is more similar to metazoan NF-κB proteins than to Rel proteins. Removal of the ANK domain allowsCo-NF-κB to enter the nucleus, bind DNA, and activate transcription. However, C-terminal processing ofCo-NF-κB is not induced by IκB kinases in human cells. OverexpressedCo-NF-κB localizes to the cytoplasm inCocells.Co-NF-κB mRNA and DNA-binding levels differ across threeCapsasporalife stages. RNA-sequencing and GO analyses identify possible gene targets ofCo-NF-κB. Three NF-κB-like proteins from the choanoflagellateAcanthoeca spectabilis(As) contain conserved Rel Homology domain sequences, but lack C-terminal ANK repeats. All threeAs-NF-κB proteins constitutively enter the nucleus of cells, but differ in their DNA-binding abilities, transcriptional activation activities, and dimerization properties. These results provide a basis for understanding the evolutionary origins of this key transcription factor and could have implications for the origins of regulated immunity in higher taxa.