Basic HTML Version
Poster Full Abstracts - Cell Biology and Signal Transduction
Poster board number is above title. The first author is the presenter
199
University of Chicago, Chicago, IL.
Eyes absent (Eya) is a conserved member of the retinal determination gene network (RDGN) and functions in multiple contexts throughout development.
In the nucleus, this protein acts as a transcriptional coactivator with its DNA-binding partner Sine oculis (So). Eya is also present in the cytoplasm, where it
functions as a protein tyrosine phosphatase. Few cytosolic Eya substrates have been discovered and little is known about its developmental importance as a
phosphatase. To identify new contexts in which Eya might function, we used multiple screening methods to search for novel interactions between it and
other proteins. Yeast two-hybrid screening uncovered many putative Eya interactors. We refined these data by evaluating the ability of RNAi-mediated
knockdown of each candidate in the eye to modify the Eya
RNAi
phenotype of adult flies. Lines that strongly enhanced or suppressed the Eya knockdown
phenotype were selected for further study. These and other previous screening efforts were hindered by the strong physical interaction between Eya and So;
little Eya is thought to exist in the cytosol in cells also expressing So (e.g. in the eye). To increase the probability of detecting interactions that occur outside
the nucleus, we simultaneously expressed Eya and knocked down candidate genes in the wing imaginal disc, which lacks significant levels of So. The
disruption of each candidate was evaluated for its ability to alter Eya levels, quantified by immunofluorescence, and to impact Eya activity, which we
inferred by measuring the size and intensity of Eya-induced ectopic Dachsund expression. This approach identified several novel interactions between Eya
and cytoplasmic proteins.
250A
Akt is Negatively Regulated by Hippo Signaling for Growth Inhibition in Drosophila.
Yaoting Deng
1
, Xin Ye
2
, Zhi-Chun Lai
1,2,3
. 1) Biochemistry and
Molecular Biology; 2) Graduate Degree Program in Genetics; 3) Department of Biology, Penn State University, University Park, PA.
Growth control of individual cells and the organs is a fundamental question in developmental biology. Hippo (Hpo) pathway functions to restrict cell
proliferation and promote cell apoptosis. However, how Hpo pathway regulates cell growth is still unclear. In our study, we found that Hpo signaling
regulates cellular growth by inhibiting akt expression through Yki inactivation. Loss of Hpo induces both Akt expression and its activity. When Hpo is
overexpressed, the outcome is opposite. We also show that Yki is sufficient to induce Akt expression. Our result suggests that Hippo signaling pathway
regulates cellular growth by repressing the Akt pathway activity.
251B
Characterization of cytoplasmic Eyes absent function in
Drosophila
eye development.
Charlene Hoi, Fangfang Jiang, Wenjun Xiong, Ilaria Rebay. Ben
May Department of Cancer Research, University of Chicago, Chicago, IL.
Understanding how information from multiple signaling pathways coordinates specific developmental programs in context-specific manners is of
particular interest. To address this, we study the dual-function transcription factor and protein tyrosine phosphatase, Eyes absent (Eya), which lies at the
center of a network of transcription factors known as the retinal determination gene network (RDGN) that is important in Drosophila eye formation. Eya’s
two functions appear to be spatially separated via a mechanism in which the non-receptor tyrosine kinase, Abelson (Abl), phosphorylates Eya to relocalize it
from the nucleus, where it regulates eye specification at the level of transcription, to the cytoplasm, where it directs morphogenetic events as a phosphatase.
We hypothesize that Abl-mediated phosphorylation of Eya facilitates interactions with cytoplasmic phosphotyrosine signaling pathways by providing
docking sites for proteins containing Src Homology 2 (SH2) and/or Phosphotyrosine binding (PTB) domains. Genetic screens of SH2/PTB domain-
containing genes identified several putative Eya interactors, which we are currently analyzing with further genetic tests and biochemistry. By elucidating the
bases of these interactions, we hope to further our understanding of Eya’s cytoplasmic function.
252C
Ligand-binding properties of the juvenile hormone receptor, Methoprene-tolerant.
Marek Jindra
1
, Jean-Philippe Charles
2
, Thomas Iwema
3
, V.
Chandana Epa
4
, Keiko Takaki
1
, Jan Rynes
1
. 1) Biology Center ASCR, Ceske Budejovice, Czech Republic; 2) Université de Bourgogne, Dijon, France; 3)
University of La Réunion, Ste Clotilde, Réunion, France; 4) CSIRO, Parkville, Victoria, Australia.
Juvenile hormone (JH) is a sesquiterpenoid of vital importance for insect metamorphosis and reproduction. The molecular basis of JH signaling remains
obscure, as a bona fide JH receptor has not yet been identified. Mounting evidence points to the bHLH-PAS protein, Methoprene-tolerant (Met), as the best
JH receptor candidate. Met was discovered in
Drosophila
through a genetic screen for resistance to the JH mimic methoprene, and was shown to bind JH. In
Drosophila
, Met acts redundantly with its paralog, encoded by the
germ-cell expressed
(
gce
) gene. Flies lacking either Met or Gce are viable and fertile;
only the double-mutants die as pupae. We have previously shown that loss of the single
Met
gene in the beetle
Tribolium
causes larvae to metamorphose
precociously, i.e. before reaching their final instar. Such a phenotype is typical for loss of JH itself, therefore supporting the idea that Met could indeed be
the elusive JH receptor. However, details of how Met transduces the hormonal signal have been missing. Here, we demonstrate that Met specifically binds
juvenile hormone (JH III) and its biologically active mimics, methoprene and pyriproxyfen, through its carboxy-terminal PAS domain. Substitutions of
individual amino acids, predicted to form a hydrophobic ligand-binding pocket, with residues possessing bulkier side chains reduce JH III binding, likely due
to steric hindrance. While a mutation that completely abolishes JH III binding does not affect a Met-Met complex that forms in the absence of methoprene, it
prevents both the ligand-dependent dissociation of the Met-Met dimer and the ligand-dependent interaction of Met with its partner bHLH-PAS protein
Taiman. These results show that Met can sense the JH signal through direct, specific binding of the hormone, thus establishing a new class of intracellular
hormone receptors.
253A
PKA upstream factors regulating autophagy in
Drosophila melanogaster
.
JUNG KIM, THOMAS NEUFELD. GENETICS, CELL BIOLOGY AND
DEVELOPMENT, UNIVERSITY OF MINNESOTA, MINNEAPOLIS, MN.
Autophagy is a self-eating process which degrades cytoplasmic organelles or proteins to produce an alternative source of nutrients under starved
conditions. Recent studies showed that cAMP-dependant protein kinase (PKA) plays a significant role in autophagy. Here, we tested G proteins, G protein
coupled receptors (GPCRs) and adenylate cyclases which are upstream factors of the PKA signaling pathway, and 12 RNAi lines of these factors affected
autophagic phenotype in Drosophila melanogaster. One of screened candidate is adipokinetic hormone receptor (AKHR), which regulates lipid and
carbohydrate homeostasis, and we hypothesize that down regulation of AKHR under starved conditions induces autophagy via PKA in D. melanogaster.