Basic HTML Version
Poster Full Abstracts - Cell Biology and Signal Transduction
Poster board number is above title. The first author is the presenter
180
VA.
Signaling by Abl kinase is important in normal cell development, differentiation, and migration. Abl protein regulates the assembly of actin, a globular
protein that is a major component of the cytoskeleton. In humans, Abl activation occurs in cells that have the Philadelphia chromosome, which carries the
mutant
Bcr-Abl
fusion gene, an oncogene that forms as a result of a translocation between chromosomes 9 and 22. As a result of this translocation, Abl
kinase activity is increased, and the structure of actin and cell migration are altered. The goal of our laboratory is to identify genes that interact with Abl
during cell migration, using
Drosophila
embryos that express Bcr-Abl as a model system. Embryos expressing Bcr-Abl die, with defects in developmental
processes that require cell migration. To identify genetic interactions with Abl, the phenotypes of embryos expressing Bcr-Abl were compared to the
phenotypes of embryos expressing Bcr-Abl and carrying a heterozygous deficiency. In this study, four deficiencies previously shown to interact with Bcr-
Abl were examined. Within each region, we tested smaller deficiencies and mutant alleles of candidate genes. For example, we dissected a region of the
second chromosome defined by two overlapping deficiencies that suppressed phenotypes associated with Bcr-Abl expression (Df(2R)ED1673 and
Df(2R)ED1715). We found that a mutant allele of
didum
, which lies in the region of overlap, suppressed Bcr-Abl associated phenotypes, while five smaller
deficiencies and mutant alleles of two other candidate genes in this region did not modify phenotypes associated with Bcr-Abl expression. We analyzed three
other interacting deficiencies in a similar fashion, and overall, we identified three genes that are likely to play a role in Abl signaling pathways. This research
was a collaborative study between Randolph-Macon College and four students and a biology teacher from Cosby High School over two summers.
175A
The integrin effectors, PINCH and RSU1, modulate actomyosin contractility in mutants of the myosin phosphatase
flapwing
via independent
mechanisms.
Julie L. Kadrmas
1,2
, Stephen M. Pronovost
2
. 1) Oncological Sci; 2) Huntsman Cancer Institute, Univ Utah, Salt Lake City, UT.
Contractility of the actomyosin cytoskeleton is essential for cell shape changes and cell migration underpinning many biological processes. Actomyosin
contraction is driven by kinase activity on serine-21 and threonine-20 of non-muscle Myosin Regulatory Light Chain, encoded by
spaghetti squash
(
sqh
).
Actomyosin relaxation requires the corresponding dephosphorylation of Sqh.
flapwing
(
flw
) encodes Protein phosphatase 1β, with the single essential
function of Sqh dephosphorylation. Strong loss-of-function mutants in
flw
have hyper-phosphorylated Sqh, with lethality due to larval muscle detachment in
a majority of animals. This phenotype of
flw
mutants overlaps with that of PINCH, a 5 LIM domain scaffolding protein encoded by
steamer duck
(
stck
).
PINCH functions to stabilize actin-integrin linkages in muscle. In
stck
mutants, embryonic lethality arises from cytoskeletal detachment from the muscle cell
membrane at sites of integrin attachment. PINCH functions as part of a stabilized protein complex that includes its direct binding partner RSU1, encoded by
ics
. We tested for a genetic interaction between
flw
and both
stck
and
ics
. We demonstrate that ectopic expression of PINCH-Flag using the native
stck
promoter fully suppresses the larval lethality of
flw
mutants, but does not reduce aberrantly high levels of phospho-Sqh as a means to promote pupariation.
Additionally, we show that elimination of RSU1 enhances the larval lethality of
flw
mutants. In the absence of RSU1,
flw
larvae exhibit an additional 3-fold
increase in levels of phospho-Sqh. Together, this supports a model in which both PINCH and RSU1 regulate actomyosin contractility in larvae, but do so via
distinct mechanisms: RSU1 can influence the phosphorylation state of Sqh, whereas PINCH acts by an alternative mechanism, perhaps by stabilizing a
constitutively contracted cytoskeleton. Future work will further define these regulatory pathways.
176B
Dissecting the regulation, interactions, and activity of the APC2-Dia complex in the formation of actin pseudocleavage furrows in the Drosophila
syncytial embryo.
Ezgi Kunttas Tatli
1
, Vince Stepanik
1
, Richa Jaiswal
2
, Bruce L. Goode
2
, Brooke M. McCartney
1
. 1) Department of Biological Sciences,
Carnegie Mellon University, Pittsburgh, PA; 2) Department of Biology and Rosenstiel Basic Medical Sciences Research Center, Brandeis University,
Waltham, Massachusetts.
Many cellular and developmental processes like cell division, cell shape change and cell migration require precise cytoskeletal rearrangements, which are
orchestrated via numerous actin and microtubule-associated proteins. The colon cancer tumor suppressor Adenomatous polyposis coli (APC) is a poorly
understood regulator of the actin cytoskeleton. One powerful model system to study the role of APC in actin organization in vivo is the Drosophila syncytial
embryo. During this stage, the embryo undergoes synchronous nuclear divisions without cytokinesis, and actin-based pseudocleavage furrows act as physical
barriers between neighboring nuclei during each division to ensure mitotic fidelity. We have previously shown that a complex between APC2 and the formin
Diaphanous (Dia) regulates the formation and extension of pseudocleavage furrows. However, the regulation of this complex, the molecular basis for their
interaction, and APC2’s affects on Dia’s actin nucleation and elongation activity are not well understood. To answer these questions, first we tested the
hypothesis that the activity of the APC2-Dia complex is regulated by APC2 phosphorylation. Drosophila APC2 is a phospho-protein and APC activity is
regulated by phosphorylation in other contexts. We show that phosphorylation of the 20 amino acid repeats (20Rs), a region known to interact with
Armadillo (Arm), plays a role in actin furrow extension. Interestingly, Dia binds to both a region of APC2 containing the 20Rs and to the SAMP repeats,
suggesting a complex interaction between APC2 and Dia that may be regulated by phosphorylation. Lastly, using in vitro assays, we are investigating the
effects of both APC2 and APC1 on the actin nucleation and elongation activity of Dia.
177C
Analysis of cell overstretching induced by microtubule depolymerization during tracheal morphogenesis.
Pierre-Marie LE DROGUEN, Antoine
GUICHET, Veronique BRODU. Institut Jacques MONOD, Paris, Paris, France.
Microtubules (MTs) are essential for many cell features such as cell shape, polarity, motility and vesicle trafficking. Through these processes, MTs are
involved in establishing epithelial structure. Notably they play a central role in directing Adherens Junction (AJ) assembly during cellularization of
Drosophila embryo. During gastrulation onwards, AJs have to be reorganized and maintained but the role of MT is poorly characterized. We have addressed
the question of MT requirement during embryonic tracheal morphogenesis. Tracheal branches arise in part from the migration of cells at the tip of branch
buds. This migration induces a pulling force on following tracheal cells as cells remain attached to each other. Hence this force triggers cell intercalation
followed by cell elongation leading to the total branch extension. Throughout this branching process, AJs are remodeled and maintained between tracheal
cells, ultimately giving rise to a branched tubular network organized around a lumen. We have established that during tracheal morphogenesis MTs are
important for the branching process as depolymerization of MT network leads to branch breaks. We first characterized more precisely this phenotype and
show that secretion of lumen products is not affected. Furthermore, the overall cell polarity seems to be maintained, and reorganization of AJ during