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Full Abstracts – CELL BIOLOGY AND CYTOSKELETON
119
1
Evidence for monomeric α-catenin as a key regulator of adherens junctions in
Drosophila
.
Ridhdhi Desai
1
, Ritu Sarpal
1
, Milena Pellikka
1
, Noboru
Ishiyama
2
, Mitsuhiko Ikura
2
, Ulrich Tepass
1
. 1) Cell & Systems Biol, Univ Toronto, Toronto, Canada; 2) Division of signaling biol, Ontario cancer Institute,
Canada.
α-catenin is a component of the cadherin-catenin complex that binds to β-catenin/Armadillo (Arm) at adherens junctions (AJs). Our analysis of recently
identified null mutations in
Drosophila
α-catenin are consistent with the conclusion that α-catenin is essential for cadherin function. To explore the
molecular mechanism of α-catenin function in vivo, we carried out a structure-function analysis of α-catenin and tested α-catenin domains in the context of
DE-cadherin::α-catenin fusion proteins. We found that the N-terminus of α-catenin, which has Arm-binding and dimerization activities, and the C-terminal
region required for F-actin binding are essential for α-catenin function. In contrast, the central region of α-catenin is a strong positive modulator that likely
undergoes secondary interactions to stabilize AJs.
As an alternative to the model that α-catenin physically links cadherin to actin, it was proposed that α-catenin acts in an allosteric manner: α-catenin is
recruited to AJs through its binding to Arm, but then detaches and forms a dimer that binds and modulates actin. To test this model we examined three
different constructs: (i) A construct which brings α-catenin dimers/oligomers to AJs without strong interactions with Arm failed to rescue a-catenin mutants
arguing against the allosteric regulation model for α-catenin. (ii) A construct that has enhanced dimerization activity reduced rescue activity and enhanced
cytoplasmic localization without any obvious enrichment on F-actin structures suggesting that α-catenin dimers are not obligatory actin binding proteins in
vivo. (iii) Finally, we show that mammalian αN-catenin does not show any dimerization activity in vitro in contrast to mammalian αE-catenin, but
nevertheless shows a strong rescue of
Drosophila
α-catenin mutants. Together, these data suggest that monomeric α-catenin acts as a physical linker between
cadherin and the actin cytoskeleton.
2
Presenilin controls kinesin-1 and dynein activity in axonal transport.
Shermali Gunawardena
1
, Ge Yang
2
, Lawrence S.B. Goldstein
3
, Kunsang Dolma
1
,
Elizabeth Spina
1
. 1) Biological Sciences, SUNY at Buffalo, Buffalo, NY; 2) 2.Lane Center for Computational Biology & Department of Biomedical
Engineering, Carnegie Mellon University, Pittsburgh, PA, 15213; 3) 3.Howard Hughes Medical Institute, Department of Cellular and Molecular Medicine
and Neuroscience, University of California, San Diego, La Jolla, CA, 92093.
Neurons and other cells require intracellular transport of essential components for viability and function. Previous work suggested that the amyloid
precursor protein (APP) can function as a kinesin-1 receptor during transport. However, how APP vesicle transport is regulated remains poorly understood.
Here we show that reduction of presenilin (PS) suppresses axonal transport defects induced by excess human APP (hAPP). However, reduction of PS
enhances axonal transport defects induced by loss of Drosophila APP (APPL). This PS-dependent enhancement is specific to APPL. No effect is seen with
reduction of PS in the context of sunday driver (syd) mutants, which disrupt transport of a different class of vesicles. Further, increased anterograde and
retrograde velocities are observed for APP vesicles in PS reduced axons, but not for synaptotagmin vesicles. These increased velocities require functional
kinesin-1 and dynein motors. Our findings suggest that PS regulates APP intracellular transport by repressing kinesin-1 and dynein motor activity perhaps by
influencing GSK3b. Thus our work has unraveled a novel function for PS in axonal transport.
3
Differential positioning of adherens junctions initiates epithelial folding during
Drosophila
gastrulation.
Yu-Chiun Wang
1,4
, Zia Khan
2,3
, Matthias
Kaschube
3
, Eric Wieschaus
1,4
. 1) Department of Molecular Biology; 2) Department of Computer Science; 3) Lewis-Sigler Institute for Integrative Genomics;
4) Howard Hughes Medical Institute, Princeton University, Princeton, NJ.
During tissue morphogenesis, simple epithelial sheets undergo folding to form complex structures. The prevailing model underlying epithelial folding
involves cell shape changes driven by Myosin-dependent apical constriction. Here we describe an alternative mechanism that requires differential
positioning of adherens junctions controlled by modulation of epithelial apical-basal polarity. Using live embryo imaging, we show that prior to the initiation
of dorsal folds during
Drosophila
gastrulation, adherens junctions shift basally in the initiating cells, but maintain their original subapical positioning in the
neighboring cells. Junctional positioning in the dorsal epithelium depends on the polarity proteins Bazooka and Par-1. In particular, the basal shift that
occurs in the initiating cells is associated with a progressive decrease in Par-1 levels. We show that uniform reduction of the activity of Bazooka or Par-1
results in uniform apical or lateral positioning of junctions and in each case dorsal fold initiation is abolished. In addition, an increase in the Bazooka/Par-1
ratio causes formation of ectopic dorsal folds. The basal shift of junctions not only alters the apical shape of the initiating cells, but also forces the lateral
membrane of the adjacent cells to bend toward the initiating cells, thereby facilitating tissue deformation. We hypothesize that in epithelial tissues in which
the levels of cortical Myosin are low and constant, junctional repositioning regulated by Par-1/Bazooka interactions may play a more prominent role to
initiate folding than does differential activation of Myosin contractility. Dorsal fold formation represents an emergent model in which the insights into this
alternative mode of epithelial folding could be further analyzed.