The Regulation of Sleep by the microRNA bantam in a Subset of Glutamatergic Neurons
Creators: name: Hobin, Michael  Patrick
Publisher: name: Brandeis University
Year: 2021
Subjects: bantam
microRNA
mushroom body
sleep

description: The sleep and wake binary is a core feature of animal life. Sleep is of deep phylogenetic origin, found throughout the metazoan lineage from cnidaria to vertebrates. This evolutionary pervasiveness suggests that sleep fulfills a vital adaptive role. The deleterious effects of sleep deprivation on behavior and physiology further illustrate the importance of sleep. Nevertheless the precise function of sleep remains a subject of considerable debate. The lack of consensus on this topic illustrates the degree to which sleep remains an enduring (and compelling) mystery. The use of genetically tractable model organisms in sleep research has provided new insights into the molecular and neuroanatomical basis of sleep function and sleep regulation. One of the discoveries to have emerged from this research is the identification of wide-spread changes in gene expression between wake and sleep states. This has been accompanied by a growing appreciation for the role of gene expression factors in the regulation of sleep.microRNAs are small non-coding RNAs that post-transcriptionally repress mRNA expression. Prior work has identified a role for microRNA in the regulation of sleep. Using Drosophila melanogaster as a model organism, we performed a systematic reverse genetics screen to identify microRNAs involved in sleep. The screen was performed using microRNA sponges to knockdown microRNA function. Among the 17 sleep-promoting microRNAs identified was bantam. Here, I follow up on the screen by characterizing the cellular, physiological and gene-regulatory basis of bantam sleep regulation.
description: I found that ban expression during development is required for normal daytime sleep. In contrast, ban expression during adulthood is required for normal nighttime sleep. I mapped the effect of bantam on nighttime sleep to a subset of glutamatergic neurons- the γ5β′2a/β′2mp/β′2mp_bilateral Mushroom Body Output Neurons (MBONs). Knockdown of ban in these neurons led to a reduction in nighttime sleep, with a pronounced effect on early night sleep. The γ5β′2a/β′2mp/β′2mp_bilateral MBONs promoted wake when activated. I demonstrated that ban acts to inhibit γ5β′2a/β′2mp/β′2mp_bilateral MBON activity. GCaMP6f imaging revealed hyperactivity in the γ5β′2a/β′2mp/β′2mp_bilateral MBONs as a result of ban knockdown. This ban knockdown-induced hyperactivity was shown to be causally related to the sleep loss phenotype. Synaptic silencing of the γ5β′2a/β′2mp/β′2mp_bilateral MBONs rescued the effect of ban knockdown on nighttime sleep. FACS sorting and RNAseq was used to identify ban-regulated target mRNAs in the γ5β′2a/β′2mp/β′2mp_bilateral MBONs. Two predicted target mRNAs (Kelch and CCHamide-2 receptor) were found to be negatively regulated by ban and to promote wake when over-expressed in the γ5β′2a/β′2mp/β′2mp_bilateral MBONs, suggesting that bantam may mediate its effects on sleep via the downregulation of these target mRNAs. These results demonstrate a novel role for bantam in the control of sleep and neural activity; and contribute to the genetic and physiological characterization of sleep regulation in an important model organism.