Natural antisense transcript of Period2, Per2AS, regulates the amplitude of the mouse circadian clock.

In mammals, a set of core clock genes form transcription-translation feedback loops to generate circadian oscillations. We and others recently identified a novel transcript at the Period2 (Per2) locus that is transcribed from the antisense strand of Per2 This transcript, Per2AS, is expressed rhythmically and antiphasic to Per2 mRNA, leading to our hypothesis that Per2AS and Per2 mutually inhibit each other's expression and form a double negative feedback loop. By perturbing the expression of Per2AS, we found that Per2AS transcription, but not transcript, represses Per2 However, Per2 does not repress Per2AS, as Per2 knockdown led to a decrease in the Per2AS level, indicating that Per2AS forms a single negative feedback loop with Per2 and maintains the level of Per2 within the oscillatory range. Per2AS also regulates the amplitude of the circadian clock, and this function cannot be solely explained through its interaction with Per2, as Per2 knockdown does not recapitulate the phenotypes of Per2AS perturbation. Overall, our data indicate that Per2AS is an important regulatory molecule in the mammalian circadian clock machinery. Our work also supports the idea that antisense transcripts of core clock genes constitute a common feature of circadian clocks, as they are found in other organisms.

Functional labeling of individualized postsynaptic neurons using optogenetics and trans-Tango in Drosophila (FLIPSOT).

A population of neurons interconnected by synapses constitutes a neural circuit, which performs specific functions upon activation. It is essential to identify both anatomical and functional entities of neural circuits to comprehend the components and processes necessary for healthy brain function and the changes that characterize brain disorders. To date, few methods are available to study these two aspects of a neural circuit simultaneously. In this study, we developed FLIPSOT, or functional labeling of individualized postsynaptic neurons using optogenetics and trans-Tango. FLIPSOT uses (1) trans-Tango to access postsynaptic neurons genetically, (2) optogenetic approaches to activate (FLIPSOTa) or inhibit (FLIPSOTi) postsynaptic neurons in a random and sparse manner, and (3) fluorescence markers tagged with optogenetic genes to visualize these neurons. Therefore, FLIPSOT allows using a presynaptic driver to identify the behavioral function of individual postsynaptic neurons. It is readily applied to identify functions of individual postsynaptic neurons and has the potential to be adapted for use in mammalian circuits.

Responses of different Drosophila species to temperature changes.

Temperature is a critical environmental variable that affects the distribution, survival and reproduction of most animals. Although temperature receptors have been identified in many animals, how these receptors respond to temperature is still unclear. Here, we describe an automated tracking method for studying the thermotactic behaviors of Drosophila larvae and adults. We built optimal experimental setups to capture behavioral recordings and analyzed them using free software, Fiji and TrackMate, which do not require programming knowledge. Then, we applied the adult thermotactic two-choice assay to examine the movement and temperature preferences of nine Drosophila species. The ability or inclination to move varied among these species and at different temperatures. Distinct species preferred various ranges of temperatures. Wild-type D. melanogaster flies avoided the warmer temperature in the warm avoidance assay and the cooler temperature in the cool avoidance assay. Conversely, D. bipectinata and D. yakuba did not avoid warm or cool temperatures in the respective assays, and D. biarmipes and D. mojavensis did not avoid the warm temperature in the warm avoidance assay. These results demonstrate that Drosophila species have different mobilities and temperature preferences, which will benefit further research in exploring molecular mechanisms of temperature responsiveness.