Prof Emi Nagoshi

Despite the progress in understanding the circadian pacemaker, the specific mechanism by which it regulates sleep remains incompletely understood. We have previously demonstrated that a substantial number of genes are rhythmically expressed in the mushroom body (MB) Kenyon cells (KCs), including Pka-C1, which encodes the catalytic subunit of protein kinase A (PKA). PKA-C1 plays a crucial role in promoting daytime wakefulness; however, the underlying mechanism remains elusive. Here, using a newly developed in vivo luciferase reporter, we show that the γ lobe is the primary site of rhythmic Pka-C1 expression. Through a combination of in silico analysis, CRISPR mutagenesis, and chromatin immunoprecipitation, we identify the transcription factor Onecut as a regulator of Pka-C1 transcriptional rhythms in γ-KCs. Furthermore, genetic trans-synaptic connectivity mapping and neuronal activity imaging reveal that the dorsal lateral clock neurons (LNds) provide inhibitory input to a subset of dopaminergic (DA) neurons in the protocerebral anterior medial (PAM) cluster, PAM-γ5, rhythmically modulating their activity. This, in turn, rhythmically activates MB γ-KCs via excitatory Dop1R signaling. Resulting γ-neuron activity rhythms drive Pka-C1 transcriptional rhythms through Onecut. Furthermore, these PKA-C1 rhythms reinforce neuronal activity rhythms, creating a feedback cycle between transcriptional and neural activity rhythms that promotes daytime wakefulness. Our findings highlight the conserved role of DA in promoting wakefulness and offer mechanistic insights into its complex regulation. More generally, this work provides a mechanistic framework for how circadian rhythms are translated into neural activity to orchestrate complex behaviors such as sleep.

Sex Peptide (SP) induces many of the most studied female post-mating responses (PMRs) in Drosophila melanogaster but has been lost multiple times in the Drosophila genus. We decided to explore the PMRs of Drosophila hydei, a species without SP. Our work shows that the PMRs in D. hydei are somewhat different than those found in D. melanogaster and may be the consequence of a selection for producing a reduced number of extremely long sperm. D. hydei females lack the substantial post-mating increase in egg production found in D. melanogaster, mostly displaying only a brief induction in the laying of stored eggs. Mated females do not show a reduction in lifespan that has been linked to changes in metabolism and egg production. To further explore the reproductive biology of this species, we performed sperm competition experiments that suggest that D. hydei females may select sperm based on characteristics linked to changes in seminal fluid proteins. This was further investigated by examining the structure of the seminal fluid-producing accessory glands and the egg laying PMRs in different Drosophila species. Finally, video-based monitoring of D. hydei females was used to uncover novel changes in circadian rhythm and light preference in mated females.

has been a pioneering model system for investigations into the genetic bases of behaviour. Studies of circadian activity were some of the first behaviours investigated in flies. The Activity Monitoring (DAM) system by TriKinetics played a key role in establishing the fundamental feedback loop of the circadian clock. Although this method has many times proven to be extremely useful, it suffers from its simplification of activity to the interruption of an infrared (IR) beam. It is blind to fly movements not disrupting the beam and any modifications to this assay to achieve better resolution often requires the purchase of new and expensive modules. We required a relatively high-throughput system to explore the potential post-mating activity changes of larger species. Rather than investing in a larger and more complex DAM system, we designed a new monitoring system that is more versatile, economic and sensitive than DAM. This new system, called DrosoVAM ( Video-assisted Activity Monitoring), is simple to implement and cost efficient, using a Raspberry Pi-controlled IR, digital video system to record multiple chambers and Python scripts that drive the deep learning software DeepLabCut, to track fly activity over multiple days.

The role of circadian clocks in regulating metabolic processes is well known; however, their impact on metabolic states across species and life stages remains largely unexplored. This study investigates the relationship between circadian rhythms and metabolic regulation in the Drosophila larval fat body, a metabolic hub analogous to the mammalian liver and adipose tissue. Surprisingly, the fat body of period null mutants, which lack a functional circadian clock in all tissues, exhibited 12-hour rhythms in gene expression, particularly those involved in peroxisome function, lipid metabolism, and oxidative stress response. These transcriptomic rhythms were aligned with 12-hour oscillations in peroxisome biogenesis and activity, reactive oxygen species levels, and lipid peroxidation. Furthermore, period mutants exhibited 12-hour rhythms in body fat storage, ultimately leading to a net reduction in body fat levels. Collectively, our results identify clock-independent ultradian rhythms in lipid metabolism that are essential for larval survival and development.

The misfolding and aggregation of TAR DNA binding protein-43 (TDP-43), leading to the formation of cytoplasmic inclusions, emerge as a key pathological feature in a spectrum of neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS) and frontotemporal lobar dementia (FTLD). TDP-43 shuttles between the nucleus and cytoplasm but forms nuclear bodies (NBs) in response to stress. These NBs partially colocalise with nuclear speckles and paraspeckles that sequester RNAs and proteins, thereby regulating many cellular functions. The laboratory of Steven Brown has recently found that the non-POU domain-containing octamer-binding protein (NONO), a component of paraspeckles, forms novel nuclear speckle-like structures in mouse cortical neurons in response to stress and sleep deprivation. These findings suggest the possibility of a functional link between NONO and TDP-43, potentially contributing to TDP-43 proteinopathy. Here, we demonstrate that pathological phenotypes caused by TDP-43 gain of function-locomotor defects and life span shortening-are exacerbated by silencing the Drosophila homolog of NONO, no on or off transient A (NonA). Additionally, NonA silencing results in an increase in nuclear TDP-43 NBs. These results provide supporting evidence for the functional link between NONO and TDP-43 and lay the foundation for dissecting underlying mechanisms.

Neurodegenerative disorders represent a leading cause of disability among the elderly population, and Parkinson's disease (PD) is the second most prevalent. Emerging evidence suggests a frequent co-occurrence of circadian disruption and PD. However, the nature of this relationship remains unclear: is circadian disruption a cause, consequence, or a parallel feature of the disease that shares the same root cause? This review seeks to address this question by highlighting and discussing clinical evidence and findings from experiments using vertebrate and invertebrate animal models. While research on causality is still in its early stages, the available data suggest reciprocal interactions between PD progression and circadian disruption.

Many organisms, including cosmopolitan drosophilids, show circadian plasticity, varying their activity with changing dawn-dusk intervals. How this behaviour evolves is unclear. Here we compare Drosophila melanogaster with Drosophila sechellia, an equatorial, ecological specialist that experiences minimal photoperiod variation, to investigate the mechanistic basis of circadian plasticity evolution. D. sechellia has lost the ability to delay its evening activity peak time under long photoperiods. Screening of circadian mutants in D. melanogaster/D. sechellia hybrids identifies a contribution of the neuropeptide pigment-dispersing factor (Pdf) to this loss. Pdf exhibits species-specific temporal expression, due in part to cis-regulatory divergence. RNA interference and rescue experiments in D. melanogaster using species-specific Pdf regulatory sequences demonstrate that modulation of this neuropeptide's expression affects the degree of behavioural plasticity. The Pdf regulatory region exhibits signals of selection in D. sechellia and across populations of D. melanogaster from different latitudes. We provide evidence that plasticity confers a selective advantage for D. melanogaster at elevated latitude, whereas D. sechellia probably suffers fitness costs through reduced copulation success outside its range. Our findings highlight this neuropeptide gene as a hotspot locus for circadian plasticity evolution that might have contributed to both D. melanogaster's global distribution and D. sechellia's specialization.

Sleep and circadian rhythm disruptions are frequent comorbidities of Parkinson's disease (PD), a disorder characterized by the progressive loss of dopaminergic (DA) neurons in the substantia nigra. However, the causal role of circadian clocks in the degenerative process remains uncertain. We demonstrated here that circadian clocks regulate the rhythmicity and magnitude of the vulnerability of DA neurons to oxidative stress in male Drosophila. Circadian pacemaker neurons are presynaptic to a subset of DA neurons and rhythmically modulate their susceptibility to degeneration. The arrhythmic period (per) gene null mutation exacerbates the age-dependent loss of DA neurons and, in combination with brief oxidative stress, causes premature animal death. These findings suggest that circadian clock disruption promotes dopaminergic neurodegeneration.

Parkinson's disease (PD) is the second most common neurodegenerative disorder, afflicting over 1% of the population of age 60 y and above. The loss of dopaminergic (DA) neurons in the substantia nigra pars compacta (SNpc) is the primary cause of its characteristic motor symptoms. Studies using and other model systems have provided much insight into the pathogenesis of PD. However, little is known why certain cell types are selectively susceptible to degeneration in PD. Here, we describe an approach to identify vulnerable subpopulations of neurons in the genetic background linked to PD in , using the split-GAL4 drivers that enable genetic manipulation of a small number of defined cell populations. We identify split-GAL4 lines that target neurons selectively vulnerable in a model of ()-linked familial PD, demonstrating the utility of this approach. We also show an unexpected caveat of the split-GAL4 system in ageing-related research: an age-dependent increase in the number of GAL4-labelled cells.

Progressive degeneration of dopaminergic (DA) neurons in the substantia nigra is a hallmark of Parkinson's disease (PD). Dysregulation of developmental transcription factors is implicated in dopaminergic neurodegeneration, but the underlying molecular mechanisms remain largely unknown. Drosophila Fer2 is a prime example of a developmental transcription factor required for the birth and maintenance of midbrain DA neurons. Using an approach combining ChIP-seq, RNA-seq, and genetic epistasis experiments with PD-linked genes, here we demonstrate that Fer2 controls a transcriptional network to maintain mitochondrial structure and function, and thus confers dopaminergic neuroprotection against genetic and oxidative insults. We further show that conditional ablation of Nato3, a mouse homolog of Fer2, in differentiated DA neurons causes mitochondrial abnormalities and locomotor impairments in aged mice. Our results reveal the essential and conserved role of Fer2 homologs in the mitochondrial maintenance of midbrain DA neurons, opening new perspectives for modeling and treating PD.