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Poster Full Abstracts - Cell Biology and Signal Transduction
Poster board number is above title. The first author is the presenter
192
the opposite phenotypes of mutation of socs36e, a negative regulator of JAK/STAT pathway. The diameter of testes of both wildtype and Upd3 mutant will
be measured at different time points. In addition, using antibody staining to mark different cell types and PH3 staining to mark dividing cells, the number of
GSCs and SSCs, as well as division rate of GSCs and SSCs will be determined as animals age. The results of these analyses will be presented.
221B
Transcriptional regulation of the
unpaired3
gene in
Drosophila
development.
Yu-Chen Tsai, Hsin-Yi Huang. Dept Life Science, Tung-hai Univ,
Taichung, Taiwan.
Unpaired3 (Upd3) is a ligand of Janus Kinase/ Signal Transducers and Activators of Transcription (Jak/STAT) signaling in
Drosophila
.
upd3
is expressed
in eye-antenna disc, gonad in the embryonic stage.
upd3
may play a role in eye, gonad, haematopoiesis and immune response. In this study, we focus on the
transcriptional regulation of the
upd3
gene and further study the upstream signaling of the
upd3
gene. We analyzed the
upd3
enhancers
in vivo
. The 22.1Kb
genomic regions around the
upd3
gene were analyzed. These
upd3
genomic fragments were cloned to enhancer-testing vector, pH-Stinger, which contains a
GFP reporter. The enhancer-testing constructs were injected into
Drosophila
and then selected for transgenic lines. The expression pattern of GFP reporter
was examined
in vivo
. We found a possible 1.2Kb
upd3
enhancer which may regulate eye, intestine cells, posterior signaling center (PSC) in lymph gland.
We further narrowed down this enhancer fragment and examine GFP pattern
in vivo
. To understand the upstream signaling of the
upd3
gene, we predicted
the possible transcriptional factor binding sites. Suppressor of Hairless (Su(H)), STAT92E and AP-1 binding sites are found in 22.1Kb genomic regions of
the
upd3
gene. We will further analyze the transcriptional regulation of the
upd3
gene in eye, gonad, and lymph gland development in
Drosophila
.
222C
Characterization of tracheal remodeling in third instar larvae through sequential imaging.
Alexandru S Denes, Oguz Kanca, Emmanuel Caussinus,
Markus Affolter. Biozentrum, University of Basel, Basel, Switzerland.
The tracheal system is an excellent model for the study of tubular organs. Little is known about late third instar and pupal stage remodeling in the tracheal
system, at least in part due to a lack of live imaging techniques. This study aims to improve our understanding of remodeling by labeling and tracking cells
within the same animal. Unlike embryos, larvae have a fully developed gut and muscular system, making imaging very difficult. Therefore, we cooled down
the animals using a custom built metal box and ice. The larvae were mounted on a slide and cover slip that fit on the box. The imaging was done with an
inverted confocal microscope. While under anesthesia, the heart continued to beat at a slow rate; other movements were essentially suppressed for the
duration of a recording session (around 20 min). Both the tracheal system and the wing disc are accessible to live imaging. Subsequently, the larvae were
transferred to food and allowed to continue development: the survival rate was close to 100% even after multiple recordings. Cell labeling was achieved
through two methods. First, we generated flies harboring a transgene encoding a nuclear localized, tandem version of mEos2 (tdEos2). Photoconverted cells
could be recognized even after multiple rounds of division. Second, we developed a version of Flybow with good spectral separation: up to five different
colors could be simultaneously recorded. Larvae expressing tdEos2 in the tracheal system were staged and individual cells in various branches of tracheal
metameres were photoconverted. We were able to show there are two temporal gradients with regard to cell division in the dorsal branches: one from the
anterior tracheomeres to the more posterior ones and another from proximal to distal with regard to the dorsal trunk. The degree of polyploidy in the dorsal
trunk along the antero-posterior axis was quantified using Flybow. Furthermore, we tracked the rounds of cell division in the second tracheomere.
223A
Computer simulation and live-imaging support a stochastic model of ventral furrow formation.
Philipp Spahn, Rolf Reuter. University of Tuebingen,
Tuebingen, Germany, Interfaculty Institute for Cell Biology, Division of Animal Genetics.
Ventral furrow formation (VFF) in Drosophila is an attractive model system to understand how an epithelium undergoes coordinated morphogenesis. In
order to initiate the internalization of ventral tissue, cells of the ventral epithelium constrict their apices leading to a furrow-like invagination into the interior
of the embryo. This apical constriction is brought about by an apically localized contracting actomyosin meshwork being tightly coupled to apically
positioned adherens junctions. It has been previously reported that the reduction of apical cell area occurs in a stereotypical incremental fashion because
phases of actomyosin contraction alternate with phases of stabilization, where the cell has to maintain its constricted state before the next contraction pulse is
set off. Here, we present a 2D computer simulation integrating a simple model of a stochastically contracting actomyosin and mechanical cell-cell coupling.
We find that rapid random contraction pulses in combination with a ventral-to-lateral gradient of contraction rate are sufficient to cause an epithelial
transformation that closely mimics the ventral furrow, as can be validated by confocal live-imaging. Unlike previously stated, we do not find apical
constriction to follow a stereotypical incremental pattern. Both simulation and live-imaging show that occasional pauses in area decrease do not need to
coincide with pauses of contraction as the cell's contraction can be compensated by contractions of neighboring cells. This can result in a temporary halt of
apical constriction or in cell stretching, both showing up as pauses in the area graph. Challenging the notion that apical constriction follows a stereotypical
pattern and only depends on the cell's own actomyosin contraction, we propose a model, in which shape change of ventral cells occurs as a stochastic process
driven by random contraction pulses and mutual mechanical interaction with neighboring cells.
224B
Candidate-based
in vivo
RNAi Screen to Identify Novel Genes Regulating Collective Border Cell Migration.
George Aranjuez
1,3
, Elizabeth Kudlaty
2
,
Jocelyn A. McDonald
3
. 1) Genetics, Case Western Reserve University, Cleveland, OH; 2) Biological Sciences, Northwestern University, Evanston, IL; 3)
Molecular Genetics, Cleveland Clinic Foundation, Cleveland, OH.
Border cells are a group of 6-10 cells that migrate during late
Drosophila
oogenesis. These cells move as a cluster that collectively follows guidance cues
to the oocyte, their migratory target. Border cell migration serves as a powerful genetic model for collective cell migration, which occurs as part of normal
embryonic development, wound healing, and tumor metastasis. To identify new genes required for collective border cell migration, we performed an
in vivo
RNAi screen to knock down genes encoding PDZ domain-containing proteins. The PDZ domain is one of the largest families of protein interaction domains
found in eukaryotes. Proteins with PDZ domains are known to regulate apical-basal polarity and signalling cascades, both of which play important roles in
border cell migration. Targeting PDZ domain-containing proteins effectively screens a larger number of genes via the protein complexes and pathways
through which the PDZ domain-containing proteins work. As a validation of our screening approach, we pulled out
baz
and
par-6
, known regulators of