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Poster Full Abstracts - Neural Development
Poster board number is above title. The first author is the presenter
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larval synapses expressing pathogenic polyQ protein or SWE also show defects in the total number of boutons, in the ratio of the pre and post synaptic size
of boutons and in the synapse length. Similar defects are observed in kinesin and dynein mutants. Abnormalities in synaptic transmission are also observed
in dynein mutants and in larvae expressing pathogenic polyQ proteins. In contrast, a synaptic protein mutant that has severe synaptic defects does not show
axonal transport defects. Together our findings suggest that problems in transport can propagate defects in synapse maturation and function; however,
synaptic problems have no direct consequence on transport. Further, proper maintenance and function of synapses requires the efficient transport of BMP
signaling components, which genetically interact with kinesin and dynein motors. Collectively our data suggest that perturbations in axonal transport can
directly contribute to synaptic abnormalities observed is degenerative diseases by affecting the transport of BMP signaling proteins.
681C
Spatial and Temporal Analysis of Axonal Transport Defects in Primary Neuronal Cultures from Drosophila Larvae.
Gary Iacobucci, Shermali
Gunawardena. Biological Sciences, State University of New York at Buffalo, Buffalo, NY.
Efficient intracellular transport is essential for healthy cellular function and structural integrity. Problems of this pathway have recently been implicated in
neuronal cell death and disease. To spatially and temporally determine how transport defects are initiated, we have developed a primary neuronal cell culture
system from Drosophila larvae to visualize, live, the movement dynamics of several cargos/organelles. In day 1 and day 2 old cultures we observe robust bi-
directional movement of six cargos/organelles. Using a MATLAB based single-particle tracker program we have analyzed these movement behaviors at
each time point. In contrast, motor protein mutant cultures show reduced movement of cargos with increased numbers of stalled blocks. Over time, these
mutant cultures also show defects in neuronal growth. Strikingly, we find that axonal blockages are not fixed, permanent blocks that impede transport as
previously thought, but are instead dynamic and can resolve over time. Under some mutant conditions blocks resolve while under others they do not. Taken
together, our results propose that non-resolving blocks may initiate deleterious pathways leading to death and degeneration while resolving blocks are
benign.
682A
The memory gene nalyot (Adf-1) functions downstream of CaMKII to regulate activity-dependent dendritic plasticity.
Christina Timmerman,
Subhabrata Sanyal. Cell Biology, Emory University, Atlanta, GA.
Understanding molecular mechanisms that underlie the complex processes of adaptive neuronal plasticity and memory is a central question in
neuroscience. In Drosophila, genetic screens have isolated several “memory mutants”, among them nalyot, a hypomorphic mutation in the transcription
factor Adf-1. Although nalyot mutants exhibit a dramatic defect in long-term memory formation, cellular phenotypes that results from a loss of Adf-1
activity remain largely unknown. Here, we show that Adf-1 plays a prominent role in the regulation of dendrite development and plasticity. Using the RP2
motor neuron as a model and in vivo 3D reconstruction techniques, we find that loss of Adf-1 inhibits basal dendrite development and precludes activity-
dependent plasticity. Adf-1 also functions downstream of CaMKII signaling since a “phospho-null” variant of Adf-1 inhibits, while a “phospho-mimic” form
of Adf-1 promotes dendrite growth. These results, together with a series of epistasis experiments, provide biological relevance for previously suspected
CaMKII-dependent phosphorylation sites in Adf-1. We also report that behavioral consequences of Adf-1 perturbation in motor neurons are severe.
Furthermore, similar manipulations in mushroom body neurons (sites of associative learning and memory) affect memory formation. Finally, to identify
novel transcriptional targets of Adf-1, we have carried out ChIP-Seq analysis from brain tissue using our anti-Adf-1 antibodies. These experiments, while
confirming the few previously known targets of Adf-1 (e.g. Alcohol dehydrogenase), have revealed high confidence binding sites upstream of neuronally
enriched genes with known functions in axon/dendrite guidance. A summary of these findings and investigations into several of these candidates will be
discussed.
683B
Comparative Analysis of larval Locomotion Activity and neuromuscular junction formation among Drosophilids.
Yunyi Yang, Mirela Belu, Claudia
Mizutani. Department of Biology, Case Western Reserve University, Cleveland, OH.
Comparative studies among related Drosophilids have the potential to reveal how functional structures, such as somatic muscles and Neuromuscular
Junctions (NMJs), may evolve across species and provide a framework to study the evolutionary basis of locomotor patterns. Results from our lab indicate
that the larval somatic body wall musculature of four Drosophila species,
D. busckii
,
D. pseudoobscura
,
D. simulans
and
D. sechellia
, display the same
stereotyped pattern of 30 muscle fibers per segment, as previously described for
D. melanogaster.
However, the number of myoblasts per individual fiber is
highly variable, resulting in fibers of small size in
D. busckii
and
D. pseudoobscura
, and large size in
D. simulans
and
D. sechellia
, in comparison to
D.
melanogaster
. Here we asked whether the increase in muscle size may have modified the development of NMJs in these species, and consequently their
locomotor patterns. By using a MatLab-based video-tracking motion analysis software, we found species-specific patterns of larval crawling movements
with significantly different rates of contractions, speed and displacement efficiency. The locomotion patterns are not directly influenced by muscle fiber size,
since species with similar muscle fiber sizes may display distinct locomotor patterns. We next analyzed the types and numbers of synaptic boutons of the
NMJ of muscle fibers 6 and 7. Our data indicate that each analyzed species has characteristic size and patterns of the NMJs, which might be responsible for
the species-specific locomotion activities. More interestingly, we have identified the presence of type II synaptic boutons in
D. sechellia
, which are not
present in fiber 6/7 of
D. melanogaster.
We are currently using this novel phenotypic variation of locomotion patterns to screen for candidate genes involved
in NMJ development based on sequence divergence within the melanogaster sub-group.
684C
The Microtubule Regulatory Protein Stathmin is Essential for Axonal Transport.
Alfredo Zuniga, Tori Pagel, Jason Duncan. Department of Biology,
Willamette University, Salem, OR, 97301.
Neurons employ a microtubule-based transport system to bidirectionally transport proteins, vesicles, and organelles between the cell body and the synaptic
terminal through the axoplasm. We have identified the protein stathmin (stai), which regulates the dynamics of the microtubule cytoskeleton, as a component
required for axonal transport in Drosophila. We have isolated several hypomorphic mutations in the stai gene that cause neuronal dysfunction resulting in
phenotypes consistent with severe defects in axonal transport. Mutant third instar larvae exhibit a posterior paralysis, or 'tail flip' phenotype, after each