Program/Abstract # 172Examining the role of the C. elegans uterine Anchor Cell in vulvamorphogenesisKathleen Estes, Wendy Hanna-RoseDepartment of Biochemistry and Molecular Biology, Pennsylvania StateUniversity, University Park, PA, USA

The Caenorhabditis elegans vulva is an excellent model system toexplore the process of organ morphogenesis. The uterine AnchorCell (AC) is vital to the coordination of development of the uterusand vulva. The AC has been well-studied for its involvement incomplex signaling events that initiate cell specification in both theuterus and the vulva. The AC is also required to form a functionaluterine–vulva connection by invading the basement membranessituated between the two organs and sitting between dorsal vulvacells, anchoring the uterus to the vulva. Recently, we have observedthat the AC plays a role in the process of vulva morphogenesis,outside of its role in cell fate specification. Using several mutants,we have shown that when the AC fails to invade the vulva, fatespecification of the vulva is largely normal, but the dorsal vulvastructure and lumen formation is perturbed. Thus, AC invasion ofthe dorsal vulva is important to establish not only the uterine–vulvaconnection but also morphogenesis of the vulva. One caveat of ourgenetic approach is that these mutants also have uterine defects, sowe are creating a system in which only the process of AC invasion isperturbed. This will allow us to understand the importance of ACinvasion on vulva morphogenesis. Additionally, we are examiningpolarity of the vulva cells and their improper cell shape tounderstand why physical contact with the AC is needed to createthe luminal structure of the dorsal vulva, but not the ventral vulva.Through the use of genetic, molecular and microscopic techniques,we hope to better understand the vital role of this cell invasionevent in vulva morphogenesis.

doi:10.1016/j.ydbio.2008.05.184

Program/Abstract # 173Notch signaling and morphogenesis of single-cell tubes in theC. elegans digestive tractJeff Rasmussen a,c, Kathryn English a,b, Jennifer Tenlen a,c,James R. Priess a,b

a Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USAb Howard Hughes Medical Institute, University of Washington, Seattle,WA 98195, USAc Molecular and Cellular Biology Program, University of Washington,Seattle, WA 98195, USA

During organogenesis of the Caenorhabditis elegans digestivesystem, epithelial cells within a cyst-like primordium develop diverseand cell type-specific shapes through largely unknown mechanisms.We here analyze the morphogenesis of two adjacent epithelial cells inthe cyst, called pm8 and vpi1, which become donut-shaped, ortoroidal, single-cell tubes. pm8 and vpi1 delaminate from the dorsalepithelium and migrate across the cyst on a transient tract of lamininbetween ventral epithelial cells, while remodeling their apicaljunctions. pm8 appears to encircle the midline by wrapping aroundfinger-like processes from neighboring cells. Finally, pm8 and vpi1self-fuse to become toroids by expressing AFF-1 and EFF-1, respec-tively, two fusogens previously shown to promote cross-fusionbetween other cell types. Notch signaling is required for theexpression of multiple genes, including aff-1, in pm8. In addition,Notch inhibits eff-1 expression in pm8, thereby preventing cross-fusion between pm8 and vpi1. Thus, the tubulogenesis of pm8 and

vpi1 involves highly orchestrated interactions with neighboringepithelial cells.

doi:10.1016/j.ydbio.2008.05.185

Program/Abstract # 174Mutagenesis screen in C. elegans suggests role of mor genes inpharyngeal developmentAndrew R. Ferrier, Pliny A. SmithDepartment of Biology, Lake Forest College, Lake Forest, IL, USA

Achieving a clear understanding of the cellular and molecularphenomena driving organ formation is one of our lab's goals. In thepresent study we conducted a mutagenesis screen using a Caenor-habditis elegans pharyngeal muscle protein, myosin-2, tagged withgreen fluorescent protein as a visual assay. The screen produced over200 pharyngeal mutant lines. Interestingly, 20 mutants manifestedshort and wide blunt pharynges, suggesting that genes required forembryonic elongation were mutated. Some blunt mutants wereviable, while others died at the L1 stage, which may reflect the degreeof defective elongation. To locate the alleles responsible for disruptingthe elongation process we performed single nucleotide polymorphism(SNP) mapping. Thus far, we successfully linked 10 different mutantphenotypes to chromosomal regions. The gene, mor-1, which resultsin a shortened, rounded pharynx, was mapped to chromosome III.Furthermore, we found another 14 similar phenotypes, which mayrepresent at least two other genes, mor-2 and mor-3. mor-2 has notbeen cloned, but is located on chromosome IV and has been shown toyield very similar phenotypes as mor-1. The mor-3 gene, a calcium/calmodulin dependent protein kinase may also be ascribed a role inabnormal pharynx development. Finally, sma-1, which is required forproper pharyngeal elongation, shares phenotypic similarity withsome of our blunt mutants. We believe that these mor genes sharethe same molecular pathway and the remaining blunt phenotypesmay be a result of defective morphogenesis. Understanding thepathway in which the mor genes work will yield a greatercomprehension of pharyngeal morphogenesis.

doi:10.1016/j.ydbio.2008.05.186

Program/Abstract # 175Overcoming genetic redundancy to identify proteins acting inC. elegans gastrulationJacob Sawyer, Trudy Li, Bob GoldsteinDepartment of Biology, University of North Carolina at Chapel Hill, USA

Morphogenesis is a critical but incompletely understood part ofanimal development. A major hurdle in the study of morphogenesis isthe redundancy of genetic mechanisms involved. Morphogenesisgenes have not fallen out of standard mutagenesis screens in diverseorganisms as often as one might anticipate, and others have proposedthat this is in part a result of genetic redundancy. Caenorhabditiselegans is an excellent model system for resolving problems ofredundancy, since several techniques allow one to simultaneouslydisrupt the function of multiple genes in C. elegans. We have tested thehypothesis of redundancy in gastrulation, the first process ofmorphogenesis in the C. elegans embryo, by combining mutantswith subtle effects on gastrulation and analyzing double mutants forsynergistic effects. Our results to date suggest that redundant genesmay contribute to the initiation of gastrulation, ingression of the E-

518 ABSTRACTS / Developmental Biology 319 (2008) 502–520

cells. With the rationale that redundant or partially redundant playersmight have subtle loss-of-function phenotypes, we conducted acandidate screen based on published microarray expression data.Our screening to date has already yielded positive results and thesegenes are being further analyzed to identify the genes that arefunctioning specifically in gastrulation.

doi:10.1016/j.ydbio.2008.05.187

Program/Abstract # 176Live imaging reveals that myoblast fusion requires dynamicremodeling of the actin cytoskeletonMary Baylies a,b, Scott Nowak a, Brian Richardson a,b

a Program in Developmental Biology, Sloan Kettering Institute, NY, NY,USAb Weill Graduate School at Cornell Medical School, NY, NY, USA

Muscle morphogenesis in both Drosophila and mammals relies onthe reiterative fusion of myoblasts. The molecular mechanisms under-lying this fusion process, however, are not well understood. Using newmethods for fixed and live imaging, we show that active remodeling ofthe actin cytoskeleton is essential for fusion in Drosophila. We haveidentified a dynamic F-actin accumulation (actin focus) that forms atthe site of fusion prior to cytoplasmic mixing. Actin focus formation isdependent on molecules that modulate myoblast adhesion. Dissolutionof the actin focus directly precedes a fusion event. Whereas severalknown fusion components localize with and regulate these actin foci,others target additional behaviors required for fusion. Mutations inkette/Nap1, an actin polymerization regulator, lead to enlarged actin focithat do not dissolve, consistent with the observed block in fusion. Weshow that Kette positively regulates SCAR/WAVE, which in turnactivates the Arp2/3 complex. Mutants in SCAR and Arp2/3 have afusion block and enlarged actin foci, suggesting that Kette-SCAR-Arp2/3participate in an actin polymerization event required for focusdissolution. We have also extended our studies to mammals, showingthat similar molecules and mechanisms are required for fusion inmouse myoblasts. Taken together, our data provide insight to the rolesand dynamics of the actin cytoskeleton and its regulators duringmyoblast fusion, leading to a revision of existing models.

doi:10.1016/j.ydbio.2008.05.188

Program/Abstract # 177Rho-kinase and myosin heavy chain are required for epithelialmorphogenesis in the Drosophila embryoRobert Simone, Stephen DiNardoDepartment of Cell and Developmental Biology, University ofPennsylvania, Philadelphia, PA, USA

Proper control and direction of epithelial morphogenesis is vital tothe reproducible generation of functional tissues. Because it is bothgenetically manipulable and easily visualized, the epidermis of theDrosophila embryo is an excellentmodel to study tissuemorphogenesis.Initially, the cells of the ventral epidermis are packed into an array ofroughly equilateral polygons. Over the course of several hours, theyundergo reproducible changes in shape and orientation and, specifi-cally, they align their anterior and posterior edges into parallel rows.Components of actomyosin contractility become enriched in thealigning epidermal domains as alignment proceeds. We hypothesizedthat actomyosin contractility may be required for proper cell alignment.

Here we also show that this alignment process is dependant on MyosinHeavy Chain/Zipper (MHC/Zip), as it is disrupted in MHC/Zip mutants.Previous studies in the Drosophila embryo have implicated Rho-kinase(Rok) in the activation of morphogenetic actomyosin contractility. Weimplicate Rok in normal cell alignment because cells fail to align whenRok function is inhibited. We suggest Myosin Regulatory Light Chain/Spaghetti Squash (MRLC/Sqh) as the relevant Rok substrate inalignment because expression of a phosphomimetic allele of MRLC/Sqh suppresses the alignment defect seen after Rok inhibition. Overall,we show that actomyosin contractility – mediated by Rok, throughMRLC/Sqh and MHC/Zip – is required for normal cell alignment in theDrosophila ventral embryonic epidermis.

doi:10.1016/j.ydbio.2008.05.189

Program/Abstract # 178The Drosophila gene dead man walking (dmw) has pleiotropiceffects on embryonic morphogenesis and the larval/pupaltransition, and interacts genetically with alphaPS3 integrin andSternopleural (Sp)Michael B. Dinkins a, James T. Warren b, Lawrence I. Gilbert b,Ellen K. LeMosy a

a Department of Cellular Biology and Anatomy, Medical College of GA,Augusta, GA, USAb Department of Biology, University of NC-Chapel Hill, USA

The Drosophila dmw gene encodes a highly conserved cathepsin B-like extracellular protein presumed to be inactive due to a substitutionof the catalytic cysteine typical of cathepsins with a serine at residue213. To explore its function, we deleted dmw by homologousrecombination and recovered a homozygous lethal allele. Developmentof dmw animals proceeds normally until the third instar stage when thelarvae undergo fat body melanization followed by delayed pupariation(24–30 h) that was not rescued by ecdysone (20E; 0.5 mg/mL) feeding.20E titers in dmw white prepuparia are not significantly reduced (n=5;pN0.2) compared to heterozygotes, though larvae failing to pupariatehave low 20E titers. Interestingly, global expression of DmwS213C-V5 indmw+ larvae produces a highly penetrant dominant negative lethalphenotype during metamorphosis with 10–20% undergoing melaniza-tion. We detect genetic interactions of dmw with alphaPS3 integrin(scab) in double homozygotes where 13.5% of embryos exhibit severegut herniation compared to a mild herniation in 4.2% of scab embryos(no phenotype in zygotic dmw embryos). Also, global expression ofDmw-V5 suppresses the dominant Sp bristle phenotype. Embryoslacking maternal Dmw fail to organize multiple tissues properly,originating with a defect in germband extension. These results suggestthat Dmw acts in the matrix to affect integrin- and perhaps wingless-mediated morphogenetic events during development.

doi:10.1016/j.ydbio.2008.05.190

Program/Abstract # 179Proteomic analysis of Drosophila fragile X mutant cleavage stageembryosKate Monzo a, Susan R. Dowd b, Jonathan S. Minden b, John C. Sisson a

a MCDB, The University of Texas at Austin, Austin, TX, USAb The Department of Biological Sciences, Carnegie Mellon University,Pittsburgh, PA, USA

Reduced fragile X mental retardation protein (FMRP) activity inbrain neurons results in the most common form of heritable mental

519ABSTRACTS / Developmental Biology 319 (2008) 502–520