Abstracts - Society for Developmental Biology

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verify the functional specificity of Mek1 and Mek2, we generated Mek1 knockin allele in which the Mek2 cDNA is under
control of Mek1 regulatory sequences. Mek1 Mek2/Mek2 homozygous knockin mice are viable, fertile and show no
apparent phenotype suggesting that both proteins are redundant and that functional difference is most likely mediated by
the expression profile. Scaffold proteins, like MP1, can modulate proteinkinase activity via the formation of a MEK1-
ERK1/2 complex. MP1 could thereforecontribute to the difference in function between MEK1 and MEK2. To clarify
therole of MP1 during development, knockout mice were generated. Interestingly, embryos Mp1-/- die between days 6 and
7 ofgestation, suggesting that MP1 is not restricted to the signaling pathway via MEK1. The analysis of Mp1-/ - phenotype
indicates a defectin the extraembryonic ectoderm but the molecular causes remain to bedetermined. Phenotypic analysis of
Mp1-/- mutants andthe study of knock-in Mek1Mek2/Mek2 will enableto provide preliminary results regarding the
functional redundancy or specificity between MEK1 and MEK2. (Supportedby CIHR)
Program/Abstract # 273
Role of ERK/MAPK pathway in syncytiotrophoblast formation during the establishment the blood-placental
barrier
Nadeau, Valerie; Charron, Jean, Centre de recherche en cancérologie de l'Université Laval, CRCHUQ, Quebec, Canada
The mammalian genome contains two ERK/MAP kinase kinase genes, Mek1 and Mek2, encoding dual-specificity kinases
responsible for ERK/MAP kinase activation. Loss of Mek1 function in mouse causes embryonic lethality, while Mek2
mutant mice survive with a normal lifespan. The Mek1 mutation causes placental defects affecting morphogenesis and
vascularization. Placental vascularization starts by the invasion of the labyrinth by the layer II of syncytiotrophoblasts
(SynT-II). This process guides the migration of the endothelial cells arising from the allantoïc tissue. Immunohistochemical
analyses demonstrate that the ERK/MAP kinase pathway is strongly activated in SynT. We have also shown that Mek1+/-
Mek2+/- mice die during gestation due to labyrinth malformations. Thus, even though Mek1 plays a predominant role in
placenta formation, Mek2 is also involved in this process. In Mek1+/-Mek2+/- mutants, the vascularization of the labyrinth
is reduced and defects in SynT-II formation lead to the accumulation of Multinucleated Trophoblasts Giant cells (MTG).
Mek1 deletion in SynT-II is not sufficient to compromise embryo survival. However, the deletion of both Mek1 allelesin
allantoïs-derived tissues in Mek1+/-Mek2+/- placenta increases the penetrance and the expressivity of the MTG placental
phenotype. Moreover, by using genetic and histopathological approaches, we have recently demonstrated that the MTGs
result from the fusion between SynT-I and SynT-II. We have also shown that the normal development of the SynT-I into a
thin layer of multinucleated cells depend on the presence of the SynT-II. Finally, Gcm1 and Pparg, ERK/MAPK targets,
are deregulated inmutant placentas and can contribute to the placental phenotype. (Supported by CIHR)
Program/Abstract # 274
A mass spectrometry-based approach to identify new interaction partners of the tyrosine phosphatase DEP-1
Walser, Michael; Hajnal, Alex, University of Zurich, Switzerland
The vulva of the nematode C. elegans serves as an excellent model to study evolutionary conserved signaling pathways
like the RAS/MAPK, DELTA/NOTCH, and WNT-pathways. Vulval development is induced by the activation of the
RAS/MAPK pathway, which specifies the 1° fate of the vulval precursorcell (VPC) P6.p. Subsequently, several negative
regulators of the RAS/MAPKsignaling pathway, such as LIP-1 and the density enhanced phosphatase DEP-1 areupregulated
in the neighboring VPCs P5.p and P7.p to inhibit the activation ofthe RAS/MAPK pathway, allowing NOTCH to
specify the 2° fate in these VPCs. In this process, the class III receptor protein tyrosine phosphatase DEP-1
negativelyregulates LET-23 EGFR signaling. Despite its importance as a tumor suppressor in many epithelial tissues,
thephysiological functions of mammalian Dep-1/Scc1 during normal development and tumorigenesis, as well as its
physiological substrates are poorly characterized. In orderto identify new interaction partners of C.elegans DEP-1, we
performed a mass spectrometry-based approach. DifferentGST-tagged versions of DEP-1 were expressed in E. coli,
affinity purified, and incubated with total C. elegans protein extracts. Proteinsthat bound to DEP-1::GST were then
identified by LC-MS/MS. Interactions of themost promising candidates were verified invitro by performing different pulldown
experiments. In addition, the biological functions of the novel identified DEP-1 interacting partners during vulval
development are currently analyzed invivo by performing RNAi experiments and genetic interaction analyses.
Program/Abstract # 275
Prioritized differentiation of stressed placental and embryonic stem cells
Rappolee, Daniel; Xie, Yufen; Slater, Jill; Puscheck, Elizabeth; Zhou, Sichang, Wayne State University, Detroit, United
States
As the mouse embryo implants into the uterusit is made of undifferentiated stem cells that will make the embryo, and
extraembryonic yolk sac and placenta. Before implantation pluripotent stem cells all divide. But within a day of