miRNA Regulation of Circadian Rhythms
Circadian clocks are present in all organisms. Twenty-four hour rhythms in behavior are driven by these molecular clocks, which can be synchronized to the environment by light. Impairment of such biological clocks is associated with sleep disorders, mood imbalance, impaired cognitive functions, dysregulated metabolism, and other disease states. Disruption of circadian rhythms can also occur as a consequence of modern life disturbances such as shift-work or jetlag. It is therefore of great interest and importance to understand the mechanisms of circadian regulation.
Drosophila has been employed as a genetic model for human clock-dependent disorders due to the strong conservation of circadian mechanisms in the two species and the simpler neural circuits of the fly. The fruit fly has ~150 clock neurons and many glial cells that contain molecular clocks composed of evolutionarily conserved proteins. Although glial cells were long thought to only provide metabolic support for neurons of the brain, research in the past 15 years has shown that a specific type of glial cell, the “star-shaped” astrocyte, performs important functions in neuronal signaling by releasing transmitters of their own. Using Drosophila as a model, the Jackson lab demonstrated that astrocytes can regulate neuronal circadian circuitry and that they are important for maintaining 24-h rhythms in behavior. When vesicle recycling is inhibited in astrocytes, for example, the flies become arrhythmic. Furthermore, the rhythmic neuronal release of the principle circadian transmitter for the fly – Pigment Dispersing Factor (PDF) – was also disrupted by this manipulation. A major aim for the lab is to identify signals released by fly astrocytes that regulate neuronal circuits and circadian behavior.
Samantha You, a PhD candidate in the Neuroscience graduate program, is working in the lab of Rob Jackson to investigate the role of glial microRNAs (miRNAs) in circadian behavior. miRNAs are small non-coding RNAs which typically bind to the 3’untranslated region (UTR) of mRNAs to inhibit translation. There is emerging literature on the function of miRNAs in circadian biology – certain miRNAs exhibit daily cycles in abundance and/or target clock mRNAs. miRNAs can target up to hundreds of RNAs, perhaps including those encoding signals used by astrocytes to regulate circadian behavior. By manipulating individual miRNAs, target mRNAs encoding putative astrocyte-to-neuron signals may be revealed.
To manipulate miRNAs, Samantha has used miRNA inhibitors termed miR-sponges. The lab of David Van Vactor and collaborators (Harvard) have created a library of miR-sponge strains that inhibit function of 145 different miRNAs. Samantha has expressed each miR-sponge selectively in glial cells and performed behavioral assays to assess the effect of miRNA inhibition on rhythmicity. From this initial screen, twenty-one miRNAs were identified as glial modulators of circadian behavior. A subgroup of these miRNAs was found to function specifically in astrocytes. To determine whether these miRNAs have a relevant physiological role, as opposed to a developmental one, we restricted miR-sponge expression in astrocytes to adulthood and identified six miRNAs that have adult physiological functions. Experiments are currently underway to identify the mRNA targets of these miRNAs; certain of them will be functionally relevant for maintaining circadian behavior. These experiments will provide additional insights about the mechanisms underlying glial regulation of circadian behavior.