are underway. Together these experiments will allow us to define the mechanism of paladin's action and understand its role in regulating neural crest cell development. Supported by NIH F32DE019973; Minnesota Medical Foundation. doi:10.1016/j.ydbio.2011.05.265 Program/Abstract # 310 Proliferation dynamics associated with cranial neural crest cell migration Dennis A. Ridenour, Rebecca McLennan, Jessica M. Teddy, Katherine W. Prather, Craig L. Semerad, Jeff Haug, Paul M. Kulesa Stowers Institute for Medical Research, Kansas City, MO, USA Neural crest (NC) cells frequently divide within the neural tube prior to exiting, but also as they migrate toward their target destinations. It is still unclear whether there is a pattern to the timing and position of NC cell proliferation along the migratory route and whether this helps to promote migration. In this study, we explored the dynamics of NC cell proliferation in the r4 migratory stream in the avian embryo. We measured the effect of division on the speed of migrating cells, the timing and position of cell divisions, and whether there was a pattern to the orientation angle of division of these cells during migration. In order to analyze NC cell pro- liferation at the population level, we measured cell cycle and pro- liferative changes between different subpopulations within a cranial (r4) migratory stream using BrdU incorporation, flow cytometry and photoactivation. We observed significant differences in proliferative capabilities between cells in the leading and trailing subpopulations. We also used immunohistochemistry to assess levels of Ki67 and cleaved caspase-3 as readouts of the number of cycling and apoptotic cells, respectively, throughout the r4 migratory stream. Our data suggests that there are differences in NC cell proliferation as a result of spatiotemporal location within the stream, and that these dif- ferences contribute to the colonization of the target destinations. doi:10.1016/j.ydbio.2011.05.266 Program/Abstract # 311 Lead and trailing cranial neural crest cells display distinct cellular and molecular profiles in response to surrounding microenvironments during migration Paul M. Kulesa a , Katherine W. Prather a , Jason M. Morrison a , Rebecca McLennan b a Stowers Institute for Medical Research, Kansas City, MO, USA b Stowers Institute for Medical Research Kulesa Lab, Kansas City, MO, USA Neural crest (NC) cell migration is an excellent model to study how cells acquire direction and maintain a migratory stream over long distances. We previously discovered evidence for NC cell chemotaxis, as well as differences in morphology and proliferation between lead and trailing NC cells, yet it is unclear when NC cells acquire direction to the chemotactic signals and how their identity is influenced by each other and the surrounding microenvironments. By analyzing NC cell nuclear orientations, we determined that NC cells establish cellular profiles along their migratory route with lower orientation at the lead and immediately after emerging from the neural tube. We next determined that lead and trailing NC cells not only differ from one another in position within the stream, but also in gene expression. We used qPCR of FACS isolated lead and trailing NC cells to examine the expression of 84 genes and found differential expression of 43 genes. When trailing NC cells were transplanted to the lead of the NC cell migratory stream, they remained at the lead of the stream, and their gene expression profile was more similar to that of lead NC cells. When trailing NC cells were ablated, remaining lead NC cells compensated for the loss of the trailing NC cells and spread out to mimic a typical NC cell migratory stream at both the morphometric and molecular levels. Our results suggest that NC cell gene expression is dependent on NC cell-cell communication and surrounding microenivonment, not on the NC cells’ origin. This analysis provides valuable insights into the mechanisms of directed NC cell migration, the behavioral and molecular differences between lead and trailing NC cells, and the plasticity of the NC cell identity. doi:10.1016/j.ydbio.2011.05.267 Program/Abstract # 312 Essential functions of the ADAM13 cytoplasmic domain in cranial neural crest cell migration Genevieve Abbruzzese a , Hélène Cousin b , Dominique Alfandari b a University of Massachusetts Molecular & Cell Biology, Amherst, MA, USA b Amherst, MA, USA ADAMs are transmembrane metalloproteases that control cell behavior by cleaving both cell adhesion and signaling molecules. The cytoplasmic domain of ADAMs can regulate the proteolytic activity by controlling the subcellular localization and/or the activation of the protease domain. We have recently shown that the cytoplasmic domain of ADAM13 is cleaved and translocates into the nucleus. Preventing this translocation renders the protein incapable of pro- moting cranial neural crest (CNC) cell migration in vivo, without affecting its proteolytic activity. In addition, the cytoplasmic domain of ADAM13 regulates the expression of multiple genes in CNC, including the protease Calpain-8. Restoring the expression of Calpain-8 is sufficient to rescue CNC migration in the absence of the ADAM13 cytoplasmic domain. Our work shows that the cytoplasmic domain of ADAM metalloproteases can perform essential functions in the nucleus of cells and may contribute substantially to the overall function of the protein. doi:10.1016/j.ydbio.2011.05.268 Program/Abstract # 313 Role of endothelin-A receptor in cardiac neural crest cell fate Yanping Zhang, Mitchell T. McKnight, L. Bruno Ruest TAMHSC-Baylor College of Dentistry, Dallas, TX, USA Congenital cardiovascular malformations are the most common birth defects affecting children but their cause generally remains unknown. Several of these defects occur in structures developing from neural crest cells (NCC). These NCC originate from the neural fold and migrate ventrally to populate the pharyngeal arches. During cardiovascular development, cardiac NCC (CNCC) participate in the asymmetric remodeling of the pharyngeal arch arteries into the great vessels and the septation of the outflow tract into the pulmonary and aortic outflows. One of the key signaling pathways regulating CNCC development involves Endothelin-A receptor (Ednra). The absence of Ednra signaling in the mouse causes severe cardiovascular defects, including persistent ductus arteriosus and coarctation of the aorta. However, the exact function of Ednra signaling in CNCC is unknown. We mapped the fate of CNCC in the cardiovascular system of the Ednra mouse and analyzed the survival and proliferation of these cells. Our data indicate that the migration of CNCC is aberrant in the cardiac outflow tract of the Ednra mutant embryos, but not in the pharyngeal arches. This migratory defect remains by E18.5 and appears to be independent of CNCC proliferation and apoptosis Abstracts 197