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Poster Full Abstracts - Neurophysiology and Behavior
Poster board number is above title. The first author is the presenter
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important for both circadian entrainment and behavioral masking, though visual input is more strongly involved in masking. To relate these studies to phase
shifts resulting from light pulses during subjective night, we exposed flies to light pulses of equal numbers of photons given over different time intervals.
Flies respond with graded phase changes to light pulses of graded intensity given over intervals from 0.1 to 360 min. Light sensitivity increases with time
interval over this range when flies are exposed to equal numbers of photons, indicating that photon input to the circadian system can integrate and store
photon information with high efficacy over many hours. We further study the dependence of this integration on visual versus cryptochrome photopigment
input.
612C
Calcium and cAMP signaling in the prothoracic gland and its role in the circadian timing of Drosophila emergence.
Angelina Palacios-Muñoz, John
Ewer. Laboratory of Neurogenetics and Development, Interdisciplinary Center of Neuroscience of Valparaíso, University of Valparaíso, Chile.
In Drosophila melanogaster the circadian clock regulates the timing of behavior and physiology. The core mechanisms that control rhythmic behaviors
consist of clock genes, which generate circadian molecular oscillations in pacemaker cells. Yet, how the activity of these genes causes outputs to be rhythmic
is poorly understood. The brain contains pacemaker neurons that regulate the circadian rhythm of locomotor activity. The prothoracic gland (PG) is a
peripheral endocrine tissue that synthesizes the molting hormones, which control metamorphosis. The PG also expresses clock genes and contributes,
together with the brain clock, to the circadian timing of adult emergence (eclosion). We investigated the pathway through which the PG causes the pattern of
Drosophila eclosion to be rhythmic by determining whether calcium or cAMP levels vary in this peripheral clock during the course of the day. To do so we
monitored calcium levels by measuring the intensity of fluorescence of a genetically-encoded calcium-sensitive GFP (GCaMP) specifically expressed in the
PG using the GAL4/UAS system. Similarly, we measured cAMP levels using the genetically-encoded cAMP sensor, Epac (Exchange protein directly
activated by cAMP). Our results show that there are consistent changes in the levels of calcium and cAMP during the course of the day. Furthermore, our
genetics studies show that the circadian rhythm of eclosion can be disrupted by interfering in the PG with the expression of clock genes and genes in the
calcium and cAMP pathways,. Thus, ours results suggest that calcium and cAMP signaling expresses a circadian rhythm and may contribute to the circadian
regulation of adult emergence.
613A
The transcription factor Mef2 is a key link between central clock, neuronal firing and the circadian regulation of axonal remodeling in Drosophila.
Anna Sivachenko, Yue Li, Katherine Abruzzi, Michael Rosbash. Dept Biol, Brandeis Univ, Waltham, MA.
The transcription factor Mef2 is a key regulator of muscle development, and mammalian Mef2 homologues regulate synaptic plasticity and neuronal
morphology. We identified Drosophila Mef2 mRNA in a screen for cycling transcripts enriched in the s-LNvs, key brain pacemaker neurons. Because these
neurons have been shown to undergo a circadian cycle of axonal remodeling, we tested the role of Mef2 and showed that it is required within the s-LNvs for
this fasciculation-defasciculation cycle as well as for activity-dependent remodeling. Moreover, the master circadian transcription complex CLK/CYC
directly regulates Mef2 transcription. ChIP-Chip analysis identified circadian binding of Mef2 to the chromatin of numerous target genes implicated in
neuronal plasticity, such as Fas2 and Fmr1. Genetic epistasis experiments showed that this regulatory hierarchy of rhythmic transcriptional regulation,
CLK/CYC->Mef2->key output genes, governs the circadian fasciculation cycle of the s-LNvs. Lastly, we showed that Mef2 is the first transcription factor
partner of the CLK/CYC complex; they synergize on a subset of CLK/CYC target genes, including the prominent circadian genes vri and Pdp1. Our results
indicate that Mef2 serves a key role for circadian output functions and possibly for the core clock itself.
614B
Identification of sex-specific transcriptome differences by RNA-sequencing.
Michelle N. Arbeitman
1
, Simon Knott
3
, Justin Fear
2
, Lauren McIntyre
2
,
Justin Dalton
1
. 1) College of Medicine, Florida State Univ, Tallahassee, FL; 2) Molecular Genetics and Mircobiology, Genetics Institute, University of
Florida, Gainesville, FL; 3) Cold Spring Harbor Laboratory, Cold Spring Harbor, NY.
In
Drosophila melanogaster
the sex determination hierarchy specifies nearly all differences in somatic tissues. This hierarchy consists of a pre-mRNA
splicing cascade that culminates in the production of sex-specific transcription factors, encoded by
doublesex
(
dsx
) and
fruitless
(
fru
). While there have been
several genomic-scale efforts to identify the genes regulated by these transcription factors, the previous techniques lacked the sensitivity and resolution to
identify many of the molecular events regulated by the sex hierarchy. RNA-sequencing (RNA-seq) technology provides an unprecedented opportunity to
identify differences in overall transcript abundance, and isoform-specific transcript abundance differences with high resolution. We have used RNA-seq to
analyze gene expression in tissues from animals mutant in the sex hierarchy genes and wild type animals. Analysis of the results identified hundreds of genes
and their isoforms regulated by this hierarchy, which had not been previously identified. Additionally, we have been using systems level approaches to gain
insight into the regulatory network downstream of
fru
, to understand how the genome is deployed to bring about sex-specific differences.
615C
Requirement of the Flightin Amino Terminal Sequence for Flight and Species-Specific Courtship Song in
Drosophila melanogaster
.
Samya
Chakravorty, Veronica Foelber, Bertrand Tanner, Jim Vigoreaux. Department of Biology, University of Vermont.
Flightin is a myosin binding protein that in
Drosophila melanogaster
is expressed exclusively in the indirect flight muscles (IFM). Flightin increases thick
filament stiffness and is essential for sarcomere stability and flight. In addition to flight, IFM is activated during male courtship song, but its precise
contribution has not been investigated. Here we show that courtship song is abolished in
fln
0
, a null mutation in flightin that also abolishes flight. Among
Drosophilids, the N-terminal flightin sequence is poorly conserved (~15% identity) compared to the rest of the protein (>70% identity). Given the role of
courtship song in mate selection and speciation, and the observation that many genes involved in sexual selection are positively selected, we hypothesize that
hypervariable flightin N-terminus influences species-specific mating song parameters. To test this, we created transgenic
D. melanogaster
strains expressing
a truncated flightin missing 62 N-terminal amino acids (
fln
Δ2-63
) and compared its flight and song properties to a wild-type rescued null control (
fln
+
).
fln
Δ2-63
is slightly flight compromised compared to
fln
+
(flight index: 3.12±0.34 vs. 4.2±0.36, respectively), but has similar tethered wingbeat frequency (~ 200 Hz)
and normal myofibrillar structure as determined by electron microscopy. Courtship song analysis showed that, compared to
fln
+
males,
fln
Δ2-63
males produce
songs with longer interpulse intervals (IPI, 60±6 ms vs. 40±1.7 ms) and higher sine song frequencies (220±3.2 Hz vs. 149±10.2 Hz). Other studies have