Abstracts - Society for Developmental Biology

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collaboration. RNA Spike-In controls from the External RNA Controls Consortium (ERCC) were used to enable
quantitative information on transcript abundance. This reference transcriptome will be avaluable resource for others
working with Nematostella and is an important first step in our high throughput Gene Regulatory Network construction
pipeline which will be widely applicable.
Program/Abstract # 263
Functional characterization of the upstream regulatory regions of XMSR, a gene involved in vascular and neural
development
Brahe, Catherine A.; Saha, Margaret, The College of William and Mary, Williamsburg, United States
XMSR, also known as APJ receptor, is a G-protein coupled receptor that functions in the development of the vascular and
nervous systems of all vertebrates. Previous studies have shown that the XMSR gene has varying levels of interspecies
conservation, both in the coding and regulatory regions. While its role in vascular development is relatively well-studied,
the role of XMSR in neural development is not well understood. XMSR is expressed within the ciliary body of the eye.
The ciliary body of Xenopus laevis is home to a large population of retinal stem cells, serving as a useful model for stem
cell research. This project aims to characterize the transcriptional regulation of XMSR in the retina. We have cloned two
kilobases of the upstream region and created a transgenic line to drive expression of GFP. Results from these experiments
indicate that this regulatory region drives expression of XMSR in the ciliary body of the retina. We have compared
expression of XMSR in these transgenic embryos to wild type embryos. Additionally we have used sequence analysis to
identify interspecies conservation of identified upstream transcriptional regulators. Using bioinformatic and transgenic
analyses of XMSR, we are conducting promoter analysis to identify the transcription factors and corresponding binding
sites responsible for the ciliary body expression and morpholino “knockdown” experiments to determine its role in
development.
Program/Abstract # 264
Genomic copy number variation during trophoblast giant cell endoreplication
Hannibal, Roberta L.; Chuong, Edward; Baker, Julie, Stanford University, United States
The placenta is a mammalian specific organ crucial for fetal development. A key feature of the placenta is a polyploid
trophoblast cell type that invades and remodels the uterus to promote flow of blood and nutrients to the fetus. In rodents,
these are called trophoblast giant cells (TGCs) and have up to 1,000N DNA content due to endoreplication. Recent work
has shown that placental specific defects in TGC endopolyploidy cause impairment to fetal growth, resulting in perinatal
death. However, the function of endopolyploidy in TGCs remains unknown. Two hypotheses are that 1) polyploidy
acquires the necessary quantity of genes while saving materials and time and 2) polyploidy regulates gene expression by
selectively amplifying certain genomic regions. We examined the genomic organization of TGCs using array comparative
genomics hybridization (aCGH). We found that certain regions of the genome are preferentially underreplicated (UR
domains). To further investigate, we compared our aCGH data to our data on RNA expression and the histone
modifications H3K27ac, H3K4me1 and H3K4me3 in cultured TGCs and their progenitors, trophoblast stem cells (TS
cells). We found that UR domains anticorrelate with RNA expression and active histone marks in both TGCs and TS cells.
As active histone marks anticorrelate with late replicating DNA in other 2N cell types, we are pursuing the hypothesis that
replication timing in TS cells causes UR domains in TGCs. Endocycles may be progressing with such speed that they do
not have time to replicate late-replicating DNA. This suggests that endocycle speed is crucial to TGC function, supporting
the hypothesis that polyploidy is important foracquiring the necessary quantity of genes while saving time.
Program/Abstract # 265
FACS-assisted deep sequencing of the zebrafish neural crest transcriptome
Hultman, Keith; LaBonne, Carole, Northwestern University, Evanston, United States
The neural crest, a vertebrate-specific population of progenitor cells, arises during neurulation and gives rise to a broad
range of derivatives including facial cartilage, bone, peripheral neurons and glia, and pigment cells. The stem cell-like
attributes of the neural crest, as well as its migratory and invasive behavior, are controlled by a network of signaling
pathways, transcription factors and effector genes referred to as the neural crest gene regulatory network (NC-GRN).
Misregulation of the NC-GRN in other cells or tissues can result in tumor progression / metastasis and other abnormalities.
We have exploited the power of next generation sequencing to identify novel components of the NC-GRN, focusing on the
zebrafish neural crest cell transcriptome. Using the Tg(Sox10:GFP) transgenic line, near pure populations of GFP positive
neural crest cells were sorted from dissociated embryos at key stages of neural crest development, and used to make cDNA
libraries for deep sequencing. This unbiased approach identified novel genes enriched in neural crest. It also found many
known neural crest GRN components, including sox10, sox9b, AP2, and snail1b, thus validating the findings and