Regulation of Dopamine Release by CASK-β Modulates Locomotor Initiation in Drosophila melanogaster

CASK is an evolutionarily conserved scaffolding protein that has roles in many cell types. In Drosophila, loss of the entire CASK gene or just the CASK- transcript causes a complex set of adult locomotor defects. In this study, we show that the motor initiation component of this phenotype is due to loss of CASK- in dopaminergic neurons and can be specifically rescued by expression of CASK- within this subset of neurons. Functional imaging demonstrates that mutation of CASK- disrupts coupling of neuronal activity to vesicle fusion. Consistent with this, locomotor initiation can be rescued by artificially driving activity in dopaminergic neurons. The molecular mechanism underlying this role of CASK- in dopaminergic neurons involves interaction with Hsc70-4, a molecular chaperone previously shown to regulate calcium-dependent vesicle fusion. These data suggest that there is a novel CASK- -dependent regulatory complex in dopaminergic neurons that serves to link activity and neurotransmitter release.Fil: Slawson, Justin B. Brandeis University; Estados UnidosFil: Kuklin, Elena A. Brandeis University; Estados UnidosFil: Mukherjee, Konark. Brandeis University; Estados UnidosFil: Pírez, Nicolas. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; Argentina. Brandeis University; Estados UnidosFil: Donelson, Nathan C. Brandeis University; Estados UnidosFil: Griffith, Leslie C. Brandeis University; Estados Unido

High-Resolution Positional Tracking for Long-Term Analysis of Drosophila Sleep and Locomotion Using the “Tracker” Program

Drosophila melanogaster has been used for decades in the study of circadian behavior, and more recently has become a popular model for the study of sleep. The classic method for monitoring fly activity involves counting the number of infrared beam crosses in individual small glass tubes. Incident recording methods such as this can measure gross locomotor activity, but they are unable to provide details about where the fly is located in space and do not detect small movements (i.e. anything less than half the enclosure size), which could lead to an overestimation of sleep and an inaccurate report of the behavior of the fly. This is especially problematic if the fly is awake, but is not moving distances that span the enclosure. Similarly, locomotor deficiencies could be incorrectly classified as sleep phenotypes. To address these issues, we have developed a locomotor tracking technique (the “Tracker” program) that records the exact location of a fly in real time. This allows for the detection of very small movements at any location within the tube. In addition to circadian locomotor activity, we are able to collect other information, such as distance, speed, food proximity, place preference, and multiple additional parameters that relate to sleep structure. Direct comparisons of incident recording and our motion tracking application using wild type and locomotor-deficient (CASK-β null) flies show that the increased temporal resolution in the data from the Tracker program can greatly affect the interpretation of the state of the fly. This is especially evident when a particular condition or genotype has strong effects on the behavior, and can provide a wealth of information previously unavailable to the investigator. The interaction of sleep with other behaviors can also be assessed directly in many cases with this method

Kepler eclipsing binary stars. VII. the catalogue of eclipsing binaries found in the entire Kepler data set

The primary Kepler Mission provided nearly continuous monitoring of ~200,000 objects with unprecedented photometric precision. We present the final catalog of eclipsing binary systems within the 105 deg2 Kepler field of view. This release incorporates the full extent of the data from the primary mission (Q0-Q17 Data Release). As a result, new systems have been added, additional false positives have been removed, ephemerides and principal parameters have been recomputed, classifications have been revised to rely on analytical models, and eclipse timing variations have been computed for each system. We identify several classes of systems including those that exhibit tertiary eclipse events, systems that show clear evidence of additional bodies, heartbeat systems, systems with changing eclipse depths, and systems exhibiting only one eclipse event over the duration of the mission. We have updated the period and galactic latitude distribution diagrams and included a catalog completeness evaluation. The total number of identified eclipsing and ellipsoidal binary systems in the Kepler field of view has increased to 2878, 1.3% of all observed Kepler targets

Central Regulation of Locomotor Behavior of Drosophila melanogaster Depends on a CASK Isoform Containing CaMK-Like and L27 Domains

Genetic causes for disturbances of locomotor behavior can be due to muscle, peripheral neuron, or central nervous system pathologies. The Drosophila melanogaster homolog of human CASK (also known as caki or camguk) is a molecular scaffold that has been postulated to have roles in both locomotion and plasticity. These conclusions are based on studies using overlapping deficiencies that largely eliminate the entire CASK locus, but contain additional chromosomal aberrations as well. More importantly, analysis of the sequenced Drosophila genome suggests the existence of multiple protein variants from the CASK locus, further complicating the interpretation of experiments using deficiency strains. In this study, we generated small deletions within the CASK gene that eliminate gene products containing the CaMK-like and L27 domains (CASK-β), but do not affect transcripts encoding the smaller forms (CASK-α), which are structurally homologous to vertebrate MPP1. These mutants have normal olfactory habituation, but exhibit a striking array of locomotor problems that includes both initiation and motor maintenance defects. Previous studies had suggested that presynaptic release defects at the neuromuscular junction in the multigene deficiency strain were the likely basis of its locomotor phenotype. The locomotor phenotype of the CASK-β mutant, however, cannot be rescued by expression of a CASK-β transgene in motor neurons. Expression in a subset of central neurons that does not include the ellipsoid body, a well-known pre-motor neuropil, provides complete rescue. Full-length CASK-β, while widely expressed in the nervous system, appears to have a unique role within central circuits that control motor output

Examples of descriptive statistics that can be gleaned from Tracker data that could not be determined from standard beam cross data (Track <i>CASK-β</i> N = 30, Track Control N = 29).

<p>Examples of descriptive statistics that can be gleaned from Tracker data that could not be determined from standard beam cross data (Track <i>CASK-β</i> N = 30, Track Control N = 29).</p

List of ANOVA results for Figure 3.

<p>The ANOVA table shows that at least one factor was significantly different for each sleep parameter. Results from individual Tukey pairwise comparisons are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037250#pone-0037250-g003" target="_blank">Figure 3</a>. All analyses were significant at DF_(3,115), <i>P</i><0.05, DAM Control (N = 30), DAM <i>CASK-β</i> (N = 30), Track Control (N = 29), Track <i>CASK-β</i> (N = 30).</p

List of ANOVA results for Figure 2.

<p>The ANOVA table shows that at there was a significant difference for each analysis type (DAM, Virtual Beam, 20%, 50%, and 100% FBL) at each time period (24, LP, or DP) except for Mean Sleep Episode Duration LP. Results from individual Tukey pairwise comparisons are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037250#pone-0037250-g002" target="_blank">Figure 2</a>. All analyses except Mean Sleep Episode Duration during LP were significant at DF_(4,30), <i>P</i><0.05, N = 7 for all analysis types.</p