also regulates Twist. Ppa binds Twists highly conserved WR domainand promotes its instability. Our findings suggest that the ubiquitin–proteasome system modulates the protein expression levels ofmultiple structurally distinct EMT regulatory factors, including Twist,Snail, Slug, and Sip1, through the actions of a common E3 ligase. TheWR domain can mediate additional protein–protein interactions thatimpact Twist function during neural crest development, distinct fromits effect on Twist stability. Together our findings provide novelinsights into the function of a key regulator of both early embryonicdevelopment and tumor progression.

doi:10.1016/j.ydbio.2011.05.420

Program/Abstract #459The role of Miz-1 in EMT and migration in the neural crestLaura K. Kerosuo, Marianne Bronner-FraserCalifornia Institute of Technology Biology, Pasadena, CA, USA

The neural crest cells undergo Epithelial to Mesenchymaltransition (EMT) as they delaminate from the neural tube andmigrate to various target destinations in the developing embryo. Byreactivation of the developmental programs due to oncogenicmisregulation of the transcriptional control mechanisms, EMT allowsthe release of mesenchymal-like cells from the parent epithelialtissue and thus leads to metastatis formation. Despite its crucialcontribution to tumor progression the gene regulatory networkbehind EMT is still relatively poorly understood. Ex ovo manipulationof the chicken embryo enables targeted in vivo knockdown of genesand real time analysis of the downstream effects. The Myc interactingzinc finger protein 1 (Miz-1/ZBTB17) is a transcription factordownstream of the TGF-b signaling pathway that promotes cell cyclearrest, epidermal stem cell adhesion and inhibits the recurrence oflymphoma formation. C-Myc directly binds to Miz-1 and repressesthe expression of its downstream target genes. Miz-1 and c-Myc areboth expressed in the delaminating and migrating chicken neuralcrest. Here, we have studied the role of Miz-1 in the regulation of theneural crest. By using the Miz-1 Morpholino, in situ hybridization andQ-PCR our preliminary data shows a significant change in theexpression pattern of several EMT and migration related genes.

doi:10.1016/j.ydbio.2011.05.421

Program/Abstract #460Transcription factors Gata4 and Gata6 play compensatory roles inpancreas developmentShouhong Xuana, Matthew Boroka, Stephen Duncanb, Lori SusselaaColumbia University, New York, NY, USAbMedical College of Wisconsin, Milwaukee, WI, USA

Gata4 and Gata6 are zinc finger transcription factors that bind to acommon DNA motif (A/T)GATA(A/G). Gata4 and Gata6 are expressedin several endoderm and mesoderm derived organs, including theliver, pancreas and heart, where they have been shown to havedistinct and overlapping functions. Gata4 and Gata6 null mice dieprior to pancreas formation, precluding the analysis of their functionsin pancreas development. To overcome this limitation, we generatedpancreas specific knock-outs of Gata4 and Gata6 using the Pdx1:Cremouse. Pancreas-specific deletion of either Gata4 or Gata6 individu-ally did not reveal obvious defects in pancreas development. To testwhether Gata4 and Gata6 may compensate for each other duringpancreas development, we generated Pdx1:Cre;Gata4fl/fl;Gata6fl/fl(G4/G6DKO) mice. The G4/G6DKO pups are hyperglycemic and themajority of mutant animals die shortly after birth. At P0, histological

and immunostaining studies demonstrated that pancreas develop-ment is severely affected, with very little identifiable exocrine andendocrine tissues. Analysis of the G4/G6DKO at e9.5 revealed thenormal induction of the pancreatic buds; however, as developmentproceeds, the pancreas displays retarded growth and reducedbranching. Molecularly, there was also reduced expression ofpancreatic progenitor markers such as Pdx1, Ptf1a, Sox9 and Nkx6.1.By e16.5, pancreas tissue in G4/G6DKO embryos is undetectable. Weare continuing to phenotypically and molecularly characterize theG4/G6DKO mice to identify the downstream developmental path-ways in the pancreas that are affected by the combined loss of Gata4and Gata6.

doi:10.1016/j.ydbio.2011.05.422

Program/Abstract # 461Dynamic expression pattern of Tbx2 and Tbx3 in the developingand adult mouse pancreasSalma Beguma, Virginia PapaioannoubaColumbia University Genetics & Development, New York, NY, USAbColumbia University, New York, NY, USA

Tbx2 and Tbx3 are closely related members of the T-box family oftranscription factors that are important regulators during normaldevelopment as well as major contributors to human developmentalsyndromes when mutated. Although there is evidence for theinvolvement of Tbx2 and Tbx3 in pancreatic cancer, so far there areno reports characterizing the expression pattern of these genes in thepancreas. In this study, we examined spatial and temporal expressionof Tbx2 and Tbx3 in mouse pancreas during development and inadult using in situ hybridization and immunohistochemistry. Ourresults show that Tbx2 and Tbx3 are expressed in the pancreaticmesenchyme throughout development beginning at embryonic day9.5. In addition, Tbx2 is expressed in epithelial-derived endocrine andductal cells during late fetal stages, during postnatal developmentand in adult pancreas. Tbx2 is also expressed in pancreaticvasculature during development. In contrast, Tbx3 is expressed inexocrine tissue in the postnatal and adult pancreas. Our resultsfurther demonstrate that Tbx2 and Tbx3 are expressed in endocrineand exocrine cell lines respectively. These dynamic changes in theexpression pattern of these transcription factors lay the foundationfor investigation of potential roles in pancreas development.

doi:10.1016/j.ydbio.2011.05.423

Program/Abstract # 462The LIM co-factor Ldb1 is enriched in pancreatic islet cells andrequired for proper cell development and functionChad Huntera, Tsadok Cohenb, Benjamin Edigerc, Crystal Wilcoxc,Shilpy Dixita, Heiner Westphald, Roland Steina, Catherine MaycaVanderbilt University Medical Center, Nashville, TN, USAbEunice Kennedy Shriver National Institute of Child Health and HumanDevelopment, Bethesda, MD, USAcChildren's Hospital of Philadelphia, Philadelphia, PA, USAdKennedy Shriver National Institute of Child Health and HumanDevelopment, Bethesda, MD, USA

The LIM-Homeodomain (HD) and LIM-only (Lmo) transcriptionalregulators play important roles in formation of many tissues andorgans. We have shown that Isl1 is the principal LIM-HD proteinexpressed in the endocrine pancreas during development and inadults. Removal of Isl1 at the onset of secondary transition in Pdx1-Crelate;Isl1Fl/Fl mice during development resulted in a reduction of

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mature endocrine cell numbers, as well as gradual loss of islet cellmass. MafA and Arx, key islet cell gene products, were also found tobe Isl1 transcriptional targets (Diabetes. 2009 Sep; 58(9):2059–69and J Biol Chem. 2011 Mar 9). The LIM domains of Isl1 can function asprotein interaction interfaces for other transcription factors and co-regulators. In this study, we investigated the impact of the LIMprotein co-factor, Ldb1, in endocrine pancreas formation. Ldb1expression was detected widely during early embryogenesis in thepancreatic buds and the surrounding mesenchyme, but later becamehighly enriched in islets and pancreatic ducts. Loss of Ldb1 in Pax6(Le)-Cre;Ldb1Fl/Fl mice was found to cause an overt reduction ininsulin+ and glucagon+ cell numbers by E18.5. The remaininginsulin+ cells appear dysfunctional, as they lacked MafA and Glut2 inbeta cells. Ldb1-deficient animals also suffered from overt diabetesdue to greatly reduced islet cell mass. Presently, we are examining bymicroarray and ChIP-Seq analysis the similarities between Ldb1 andIsl1 regulation of beta cells, with the expectation that differences maybe found due to the expression Lmo2 and Lmo4 in the developingpancreas.

doi:10.1016/j.ydbio.2011.05.424

Program/Abstract #463Vangl2, aPKC and VAMP1; the interactions of polarity proteinswith trafficking vesicle proteins in the Xenopus oocyteSang-Wook Cha, Emmanuel Tadjuidje, Christopher Wylie,Janet HeasmanCCHMC, Cincinnati, OH, USA

The Xenopus oocyte is a polarized cell with a distinct animal/vegetal axis. It contains the components of both PCP and apical /basalpolarity systems, including Vangl2 and flamingo (planar cell polarityproteins) and aPKC, PAR3 and 6 (apical basal polarity proteins). Tobegin to address the relative roles of these systems in the oocyte, weexamined the localization and function of Vangl2 and aPKC. We foundthat: 1. Vangl2 is enriched in radially arranged islands in the animalhemisphere. 2. Vangl2 co-localizes and physically interacts withVAMP1, a component of post-Golgi membrane trafficking vesicles. 3.The localization of animally localized VAMP islands depends uponVangl2 protein. 4. The arrangement of VAMP1/Vangl2 complexesdepends on the stable acetylated microtubule cytoskeleton 5. aPKCalso physically interacts with Vangl2 6. aPKC is required for thestability of the radially arranged acetylated microtubule cytoskeletonand the location of VAMP1/Vangl2 complexes. We conclude that bothmaternal aPKC and Vangl2 are essential for the polarity of theXenopus oocyte.

doi:10.1016/j.ydbio.2011.05.425

Program/Abstract #464A novel role for a Cdc42 effector protein in Xenopus neurogenesisAlissa Hulstranda, Douglas Houstonb

aUniversity of Iowa Department of Biology, Iowa City, IA, USAbUniversity of Iowa, Iowa City, IA, USA

Many developmental events, such as axis determination and cellmovement, are dependent on the asymmetrical distribution ofproteins and RNAs. In Xenopus laevis, several maternal mRNAsessential for normal development are localized to the oocyte vegetalcortex. In this work we characterize a novel cortex-enrichedtranscript, cdc42 effector protein 4-like (cep4l). CEPs bind cdc42and related small GTPases, which regulate many cellular functions.cep4l is expressed in the oocyte vegetal cortex and throughout

embryonic development, including expression in migratory cellsduring gastrulation, neural crest in neurulae and tailbud stages, andneural regions in older embryos. Misexpressed cep4l RNA causesconvergent extension defects and induces ectopic neuronal markerexpression, indicating a role in neurogenesis. Experiments to identifyupstream and downstream pathways indicate roles for FGF as well ascdc42. Co-expression studies with another neuronal inducer, FGF8a,demonstrate an enhancement of cdc42 binding and ectopic neuro-genesis. The effects of both cep4l and FGF8a are independent ofproliferation. We also present loss of function data showing a rolefor cep4l in normal axial and nervous system development, as wellas a requirement for FGF8a-induced neurogenesis. Although theroles of small GTPases in cell division, migration, and adhesion arewell-characterized, our results suggest novel roles and pathways forthese proteins and their effectors in neural fate patterning andneurogenesis.

doi:10.1016/j.ydbio.2011.05.426

Program/Abstract #465Expression in dorsal–lateral regions of Drosophila early embryosis supported by Grainyhead-mediated anti-repressionMayra Garcia, Angelike StathopolousCaltech, Pasadena, CA, USA

The Drosophila pre-gastrula embryo is patterned by a nucleargradient of the transcription factor Dorsal, which supports expressionof genes in defined domains along the dorsal–ventral axis. Currentmodels postulate that limiting amounts of Dorsal establish the dorsalboundaries of gene expression. In the case of the gene intermediateneuroblast defective (ind) which is expressed in a dorsal–lateralstripe, in addition, EGFR signaling also supports ind expression. Wehave evidence suggesting repressors are necessary for the sharpdorsal border of ind. A synthetic enhancer analysis of the indenhancer located a short 12 base pair repetitive sequence (“A-box”)that mediates transcriptional repression in dorsal regions of embryos(Stathopoulos & Levine, DevBio 2005). We found that this elementalone is sufficient to mediate repression in dorsal regions, further-more, when this element is mutated in the full-length enhancerexpression is expanded dorsally. We identified proteins that bind thiselement using affinity chromatography and mass spectrometry. Oneof the factors we identified is grainy head (grh), a DNA-bindingprotein, which surprisingly we found acts as an activator to supportexpression of ind and has been shown to exhibit context-depedentactivation that is influenced by MAPK signaling. We believe that grhacts to inhibit the repressor that sets the dorsal border of ind bycompetitively binding to the A-box element. Instead of limitingactivators being responsible for establishing the dorsal boundary ofind, we propose instead that expression of this gene is supported bythe grh activator that functions to limit the effects of repressors.

doi:10.1016/j.ydbio.2011.05.427

Program/Abstract # 466A high throughout sequencing-based screen for sea urchinskeletal patterning genesArlene Reynaa, Hajerah Hammeduddina, Christy Lia, Evan Bardota,David Leea, Finnegan Hewitta, Michael Piacentinoa, Patrick Ferrella,James Chaveza, Amanda Corea, Jasmin Coulombe-Huntingtona, AlbertPoustkab, Cynthia A. Bradhamc

aBoston, MA, USAbBerlin, GermanycBoston University Biology, Boston, MA, USA

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