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Poster Full Abstracts - Cell Biology and Signal Transduction
Poster board number is above title. The first author is the presenter
203
266B
Identifying novel nuclear targets for MAPK/Erk2.
Rona Grossman
1
, Tatyana Shestkin
1
, David Engelberg
2
, Gerardo Jiménez
3
, Ze'ev Paroush
1
. 1)
Developmental Biology and Cancer Research, IMRIC, Faculty of Medicine, The Hebrew University, Jerusalem, Israel; 2) Biological Chemistry, The
Alexander Silberman Institute of Life Sciences, The Hebrew University, Jerusalem, Israel; 3) Institut de Biologia Molecular de Barcelona-CSIC and
Institució Catalana de Recerca i Estudis Avançats, Barcelona, Spain.
Receptor Tyrosine Kinase (RTK) signaling pathways control key cellular processes such as proliferation, migration and cell fate specification, in normal
development and in disease. RTK pathways signal through an intracellular cascade of kinases, culminating in the phosphorylation and activation of the
downstream effector kinase, MAPK/Erk2. Once active, MAPK/Erk2 enters the nucleus where it phosphorylates transcriptional regulators, thereby modifying
their function. This brings about coordinated changes in gene expression profiles that are imperative for subsequent cellular decisions. Despite the critical
roles played by RTK pathways in various developmental processes, to date only a few confirmed transcription factors that are directly targeted by
Drosophila
MAPK/Erk2 are known. We have, therefore, established a genome-wide proteomics screen in order to uncover new direct nuclear MAPK/Erk2
substrates. So far, our assay has identified 35 putative targets for MAPK/Erk2, some of which have been previously reported (e.g., Bicoid). For several
newly selected MAPK/Erk2 targets, we have generated transgenic flies expressing unphosphorylatable as well as phosphomimetic derivatives. This
approach will help validate bona fide MAPK/Erk2-regulated targets, since expression of these variants is expected to exert differential outcomes in vivo.
Recognizing novel MAPK/Erk2 substrates should ultimately enhance our understanding of how RTK signaling pathways induce specific cellular responses
during development.
267C
In vivo
analysis of the Midkine/Pleiotrophin fly homologues Miple1 and Miple2.
Fredrik Hugosson
1
, Camilla Sjögren
1
, Ludmilla Hedlund
1
, Anna Birvé
2
,
Ruth H Palmer
1
. 1) Department of Molecular Biology, Umeå University, Umeå, Sweden; 2) Department of Medical Bioscience, Umeå University, Umeå,
Sweden.
The growth factors Midkine (MDK) and Pleiotrophin (PTN) form a family of Heparin binding proteins that have anti-apoptotic, angiogenic, mitogenic,
chemoctactic and transforming activity. Midkine and Pleiotrophin have separately been reported as candidate ligands for the Receptor Tyrosine Kinase
(RTK) Anaplastic Lymphoma Kinase (ALK)
in vitro
and with this in mind we set out analyse the
Drosophila
MDK/PTN homologues, named
miple1
and
miple2
and their possible role as functional ligands for the fly ALK receptor
in vivo
.
In situ
analysis show that the two genes have a interesting
complementary expression to ALK during embryogenesis, with
miple1
in developing CNS and
miple2
in the endoderm. Enhancer analysis using the
UAS/GAL4 system show a CNS specific expression of
miple1
in larvae and adults and a broad expression of
miple2
in mouth region,trachea ,gut and brain
in larvae and adults. To analyse the function of
miple1
and
miple2 in vivo
, we have generated single deletion mutants, they are homozygous viable and lack
obvious developmental defects. There is a possibility of a redundant function so we in next step generated
miple
double mutants. Unexpectedly, they also are
viable. Interestingly, in over-expression experiments can Miple1 and Miple2 rescue the gut phenotype seen in mutants for the bonafide ALK ligand Jelly
Belly (Jeb), and this result is dependent on ALK activity, suggesting that in some biological context can they function as ligands for ALK. We are currently
analysing developmental defects seen during embryogenesis and also the effect on lack of Miple proteins in the adult fly, with focus on gut and brain
function.
268A
Dynamic regulation of the transcriptional repressor Capicua by localized receptor tyrosine kinase signaling.
Victoria M. Sanchez
1
, Oliver Grimm
2
,
Yoosik Kim
1
, Jordi Casanova
3
, Eric Wieschaus
2
, Stas Shvartsman
1
. 1) Chemical and Biological Engineering, Princeton University, Princeton, NJ; 2)
Department of Molecular Biology, Princeton University,Princeton, NJ; 3) Institut de Biologia Molecular de Barcelona, Parc Cientific de Barcelona, Spain.
Receptor tyrosine kinases (RTKs), signaling through MAPK, control a wide range of biological processes, in many cases through regulation of
transcription. In one mode of regulation, nuclear levels of the HMG box transcriptional repressor Capicua are reduced by RTK signaling. However, the
mechanism of Capicua down-regulation by MAPK is not well understood. Making use of photoswitchable and fluorescently tagged Capicua constructs, we
characterize the temporal dynamics of the nuclear Capicua gradient, as well as the effect of RTK signaling on its nucleocytoplasmic shuttling properties in
the
Drosophila
embryo. Based on these results, we propose a biophysical model which contains a mobile and an immobile pool of Capicua. In this model,
RTK activation regulates Capicua by primarily affecting the nuclear import and export rates of the mobile pool. Furthermore, this effect alone is sufficient to
explain the formation and temporal evolution of the observed Capicua nuclear gradient in the terminal patterning of the embryo.
269B
Regulation of midgut metamorphosis via coordinated action between receptor tyrosine phosphatase Ptp52F and TER94/VCP.
Abirami
Santhanam
1,2,3
, Guang-Chao Chen
1,2,3
, Tzu-Ching Meng
1,2,3
. 1) Institute of Biological Chemistry, Academia Sinica, Taipei, Taiwan; 2) Institute of
Biochemical Sciences, National Taiwan University, Taipei, Taiwan; 3) Taiwan International Graduate Program, Academia Sinica, Taipei, Taiwan.
In
Drosophila
, a number of cellular processes including proliferation and differentiation are regulated by protein tyrosine phosphatases (PTPs).However, to
date the mechanisms by which PTPs regulate the developmental processes remains elusive especially in case of receptor PTPs (RPTPs) which are majorly
attributed to the regulation of axon guidance and synaptogenesis decisions in
Drosophila
embryos and larvae. To reveal the other potential functions we
utilized systematic data mining approaches focusing on RPTP expression profiles during critical stages of development. This lead to the identification of a
highly midgut enriched RPTP-the PTP52F especially in the larva-pupa transition during which the ecdysone action kicks in. Results from real-time PCR and
cell based experiments confirmed RPTP52F as an ecdysone response gene. Genetic studies showed a critical role of PTP52F in midgut metamorphosis
during larva pupa transition. Using a substrate-trapping strategy we identified, transitional endoplasmic reticulum ATPase94 (TER94), ortholog of human
Valosin Containing Protein (VCP) as a bonafide substrate of PTP52F. Interestingly, tyrosine 800 of TER94 which is phosphorylated by Src kinase is
targeted and dephosphorylated by PTP52F. We showed that PTP52F mediated dephosphorylation of TER94 could facilitate the ubiquitin mediated
degradation of various proteins including
Drosophila
inhibitor of apoptosis1 (DIAP1) a key regulator controlling midgut cell death. In vivo evidences
demonstrated that the forced expression of TER94 rescued the defect of midgut metamorphosis induced by knockdown of PTP52F, suggesting the