Basic HTML Version
Poster Full Abstracts - Cell Biology and Signal Transduction
Poster board number is above title. The first author is the presenter
190
epithelial polarity. We are currently working to assess the specificity and mechanism of Moesin's interaction with Crumbs and its role in trafficking, as well
as to probe the role of Crumbs trafficking at particular stages of oogenesis. .
213C
The chocolate and maroon genes are involved in intracellular transport.
Rishi Singhal
1
, Aminah Wali
1,2
, Paaqua Grant
1
, Diana Johnson
1
. 1) Biological
Sciences, George Washington University, Washington, DC; 2) University of Maryland, Baltimore County.
There are over one hundred genes that affect eye color in Drosophila. A number of them produce products that function in intracellular transport. We used
deletion analysis to find candidate genes for both chocolate (cho) and maroon (ma). We used sequencing and transgenic rescue experiments to identify and
confirm their gene identities. The cho gene is a vacuolar ATPase, CG2934. The gene has previously been identified as playing a role in Notch signaling and
vacuolar acidification. It is expressed at moderate to high levels throughout development. It is also very highly expressed in many adult tissues including the
gut, Malpighian tubules and is highly expressed in the brain, head and eye. The ma gene is a vacuolar protein sorting gene, CG8454, Vps16. It was
previously shown to be required for trafficking to lysosomes and for the development of pigment granules. It is also expressed throughout development,
though at lower levels than the cho gene. Gene expression of cho is at its highest at the late embryo and early larval stages. Expression of ma peaks later in
the late larval to mid pupal stages. Like cho, ma's adult expression is at relatively high levels in the brain, head, eye, gut, and Malpighian tubule. Knockouts
of both genes have been reported to be lethal. The cho mutant allele contains one base change resulting in a missense mutation while the ma allele shows a
deletion of nine bases causing a deletion of three neighboring amino acids. Interactions between these alleles and other trafficking genes are being
investigated to determine whether the mutant alleles' effects are limited to development of pigment granules or also affect other functions of the proteins.
214A
PP2A binds to β
H
-spectrin and modulates endosomal maturation and
zonula adherens
stability.
Graham Thomas
1
, Seung-Kyu Lee
1
, Elizabeth Klipfell
2
,
Joanna Sandilos
1
. 1) Dept Biology/BMB, Penn State, University Park, PA; 2) Cleveland Clinic, Cleveland, OH.
β
H
-spectrin (β
H
) in conjunction with α-spectrin forms the apical membrane skeleton, an F-actin-based cytoskeletal meshwork associated with the apical
plasma membrane. β
H
(
karst
) mutants result in perturbation of the
zonula adherens
(ZA) without a conspicuous loss of polarity. We have shown that β
H
is
recruited to the apical membrane
via
the FERM-binding domain of the apical polarity determinant Crumbs, a region that has a specific role in stabilizing the
ZA. Multiple lines of evidence suggest that β
H
modulates endosomal trafficking, and our current model suggests that β
H
remains associated with internalizing
vesicles/endosomes until it is released by Annexin B9 at the multivesicular body (MVB) stage. Annexin B9 is required for efficient trafficking through the
MVB and to maintain a high-fidelity apical-lateral boundary.
D
E-Cadherin is amongst the cargoes in this pathway. The C-terminal segment 33 of β
H
(βH33)
appears to be a focal point for these trafficking activities: Overexpression of this domain disrupts endocytosis leading to membrane expansion and is the
binding site for Annexin B9. Here we report that βH33 binds to the dual specificity protein phosphatase PP2A, which has demonstrated roles in the
regulation of apicobasal polarity. Through knockdown and overexpression experiments and we provide evidence that PP2A sustains the ZA. In addition,
levels of the MVB markers Hrs and Vps16 are altered in a tissue specific fashion when PP2A is overexpressed or knocked down. This suggests that the
effects of PP2A on the ZA may arise through modulation of the MVB and the recycling of ZA components in the β
H
/Annexin B9 pathway. We also
demonstrate that PP2A modulation dramatically affects the distribution of cellular F-actin. We hypothesize that Crumbs recruits β
H
to act as a scaffold for
PP2A to maintain ZA itegrity
via
appropriate recycling of
D
E-Cadherin.
215B
V0-ATPase subunit a1 regulates vesicle sorting through binding to t-SNARE acceptor complexes.
Dong Wang
1,4
, W.Ryan Williamson
1,4
,
Sankaranarayanan Srinivasan
2
, Daniel Epstein
1
, Florante A Quiocho
2
, P. Robin Hiesinger
1,3
. 1) Dept Physiology, UT Southwestern Med Ctr, Dallas, Texas
75390; 2) Verna and Marrs McLean Department of Biochemistry and Molecular Biology Baylor College of Medicine, Houston, Texas 77030; 3) Green
Center Division for Systems Biology, University of Texas Southwestern Med Ctr, Dallas, Texas 75390; 4) Equal contribution.
The vesicular ATPase is a multi-subunit proton pump whose insertion into specific target membranes is determined by different V0a subunits. Here we
show that a SNARE-binding deficient variant of the neuronal V0a subunit V100 fails to restore neurotransmission and reveals intracellular vesicle sorting
defects. In neurons, V100 binds to the target SNAREs Syntaxin and SNAP25 under exclusion of the vesicle SNARE Synaptobrevin. However, the V100 N-
terminus does not bind to the exposed SNARE domain of ‘open’ Syntaxin, but binds to the helical bundles from by ‘closed’ Syntaxin or a Syntaxin/SNAP-
25 acceptor complex. Binding to this t-SNARE acceptor complex is reduced when Synaptobrevin is present. V100 has a low t-SNARE binding affinity and
restricts n-Syb function in vivo, suggesting a function prior to v-/t-SNARE complex formation. We propose that the SNARE interaction of V100 exerts an
intracellular sorting function and show that the V100 N-terminus fused to the C-terminus of the v-SNARE Synaptobrevin is sufficient to execute this
function in vivo.
216C
The interaction between two JAK signaling ligands: Upd and Upd3.
Qian Chen, Douglas Harrison. Dept Biol, Univ Kentucky, Lexington, KY.
JAK/STAT signaling pathway is highly conserved between mammals and
Drosophila
. But unlike the mammalian JAK signaling, which can be stimulated
by a variety of cytokines and growth factors, the
Drosophila
JAK signaling has only three ligands identified: Upd, Upd2 and Upd3. Upd is the first ligand
identified in this pathway and it is so far considered to be the most essential one. While Upd2 and Upd3 are also shown to participate in the fine-tuning of the
signaling, how they cooperatively regulate the pathway with Upd is not fully understood. The expression patterns of
upd2
and
upd3
overlap with that of upd
during several developmental processes.
upd2
and
upd
express in identical stripes in embryos, while
upd3
and
upd
co-expressed in the polar cells of egg
chambers and posterior region of eye discs. Given the overlapping expression pattern, we hypothesize that the three ligands cooperatively regulate by
forming different ligand complexes. Here we show homotypic and heterotypic interaction between Upd and Upd3 using Biomolecular Fluorescence
Complementation (BiFC), and the results were confirmed by yeast two-hybrid and co-immunoprecipitation. To determine the sequence that is essential for
the physical interaction, we aligned the three ligands and identified six short consensus domains that are distributed across the proteins. Each of the
consensus sequences on Upd3 was substituted by five Alanine residues individually. Each Upd3 substitution variant is being tested for interaction with intact
Upd and Upd3 by BiFC. Finally, the signaling intensity stimulated by different ligand complexes and the upd3 substitution variants will be compared by a