Abstracts - Society for Developmental Biology

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ROS levels. Collectively, our results elucidate the importance of the crosstalk between EGFR/Ras and RBF1 in
coordinating cell cycle progression and survival.
Program/Abstract # 402
An investigation of Blastoderm specific gene 25D, a potential pole cell specifying gene.
Kowanda, Michelle A.; Yee, Stephanie; Liu, Niankun, McGill University, Montreal, Canada; Lécuyer, Eric (Institut de
Recherches Cliniques de Montréal, Montreal, Canada); Lasko, Paul (McGill University, Montreal, Canada)
Specific localized mRNAsare essential for germline specification in Drosophila melanogaster. These mRNAs accumulate
in the posterior pole plasm of the early embryo and are takenup by the pole cells, or primordial germ cells. While
numerous mRNAs have beenfound to localize at the posterior of the early embryo, the functions of manyin germ cell
development have not been determined. Additionally, a mechanism of active transport has been found to localize a subset
of germ plasm mRNAs to thepole cells. Active transport along astral microtubules creates a feature called‘RNA islands’ in
stage 3 embryos, ensuring that specific mRNAs are incorporated into the primordial germ cells. Interestingly, many wellcharacterized
mRNAs that perform crucial roles in pole cell development are recruited to RNA islands, such as germ cellless
and arrest. Blastoderm specific gene 25D (Bsg25D) mRNA also localizes to RNA islands but its function has not been
characterized. We have found that Bsg25D protein localizes to the posterior of region 2b of the germarium and posterior of
the oocyte during stages 1-10 of oogenesis. Bsg25D translation is reduced in homozygous mutant vasa 1 /vasa 1 ovaries,
although Bsg25D mRNA is localized to the oocyte throughout stages 2-10 of oogenesis. Finally, Vasa binds in vitro to the
3’UTR of Bsg25D containing a U-rich motif similar to the Vasa-binding site in mei-P26 mRNA. Thus far, our analysis
suggests that Bsg25D is a target of Vasa-mediated translationalactivation.
Program/Abstract # 403
Identification of a conserved motif in mRNAs that localize to RNA islands during Drosophila embryogenesis
Yee, Stephanie; Kowanda, Michelle, McGill University Biology, Montreal, Canada; Li, Xiao; Morris, Quaid; Lipshitz,
Howard (University of Toronto, Toronto, Canada); Lecuyer, Eric (Institut de Recherches Cliniques de Montreal,
Montreal, Canada); Lasko, Paul (McGill University Biology, Montreal, Canada)
During early Drosophila embryogenesis, post-transcriptional control of gene expression is essential for directing
development. The posterior localization of maternal transcripts to the pole cells of the embryo establishes a molecular
asymmetry, which is necessary for germ line specification. According to two ongoing large-scale in situ hybridization
screens performed on Drosophila melanogaster embryos, a small fraction of maternally contributed mRNAs localize to the
pole cells (Lecuyer et al., 2007; Tomancak et al., 2007). The posterior localization of those transcripts that later enrich in
the pole cells can be divided into three distinct categories: 1) localization to the pole plasm during oogenesis, 2)
enrichment to the pole plasm followed by formation of RNA islands around pole cell nuclei in the stage 3 embryo, and 3)
accumulation in the pole cells by stage 4. Using fluorescent in situ hybridization, we found that most mRNAs that
accumulate in D. melanogaster pole cells also do so in D. simulans and D. virilis. We have used bioinformatics tools to
identify a consensus motif within the 3’UTR of D. melanogaster mRNAs that localize to RNA islands. The functional
significance of the putative motif is currently being tested by expressing transgenes with the motif disrupted in the 3’UTRs
of gcl, gwl and pgc , three mRNAs that contain the motif, and assessing for defects in RNA localization. Altogether, our
studies will provide further mechanistic insight into germ cell development through the identification of conserved
sequences and structural motifs.
Program/Abstract # 404
A structure-function study of Vasa in Drosophila early development
Dehghani, Mehrnoush; Lasko, Paul, McGill University, Montreal, Canada
Vasa is a DEAD-box RNA-binding protein essential for germ cell specification in a variety of organisms. DEAD-box
helicases, including Vasa, were previously believed to be non-sequence specific, yet Vasa is known to regulate translation
of specific mRNAs. Previous work in our lab (Liu et al. 2009) indicated that Vasa can distinguish a (U)-rich motif in the
3’UTR of its mRNA targets such as mei-P26, and that this specificity is conferred by the part of Vasa sequence residing
outside of the motifs conserved among DEAD-box proteins. We hypothesize that the sequence-specific binding is
associated with one or more RGG motifs in the N-terminus of Vasa, since these sequences act as auxiliary motifs that
confer specificity to some other RNA binding proteins. To investigate this, we generated gfp-vasa constructs encoding
proteins with progressive deletions in their N-termini to eliminate different numbers of RGG domains. These were
expressed in vasa-null flies to study their localization pattern as well as their ability to rescue different aspects of Vasa
function. Our initial data suggest that the RGG motifs in Vasa sequence act redundantly. Furthermore, proteins lacking
more than six RGG motifs can still partially restore Vasa function; however, expressing such proteins at a high level