A Study on the Effect of YEATS2 Gene Knockdown on Dopaminergic Synaptic Integrity and Epileptic-like Behavior in Drosophila

Background Introduction

Epilepsy is a common neurological disorder characterized by abnormal electrical activities in the brain. These abnormal activities can lead to seizures and other neurological symptoms. Familial adult myoclonic epilepsy (FAME) is a rare autosomal dominant inherited disease characterized by cortical myoclonus and sporadic seizures. There are currently six known types of FAME (FAME1-FAME6), each associated with pentanucleotide repeat expansions in different genes. Specifically, FAME4 is caused by the expansion of TTTTA/TTTCA repeats located in the first intron of the YEATS2 gene. While studies suggest that these repeat expansions exert their effects primarily through RNA toxicity, the exact function of the affected genes in neurons remains unclear.

Source of the Paper

This paper was published online on December 20, 2023, in the journal “Progress in Neurobiology.” The study was conducted by Luca Lo Piccolo and several other scholars from research centers including Chiang Mai University, the University of Konstanz, the Max Planck Institute of Animal Behavior, Khon Kaen University, and Chulalongkorn University.

Detailed Research Process

Experimental Design and Methods

The study used Drosophila as a model organism to explore the impact of knocking down the YEATS2 gene (referred to as dYEATS2) on neural function and behavior. The specific experimental procedures were as follows:

1. Drosophila Strains and Culturing: Drosophila were reared on a standard cornmeal yeast glucose medium under conditions of 25°C temperature, 55% humidity, and a 12:12 light-dark cycle.

2. Gene Knockdown: The Gal4/UAS system was employed to knock down dYEATS2 expression using two independent RNA interference (RNAi) sequences (dYEATS2 ir-1 and dYEATS2 ir-2). The experiments utilized elavC155-Gal4 and nSyb-Gal4 drivers to express RNAi in all neurons during larval and adult stages, respectively.

3. Behavioral Experiments: Experiments include testing Drosophila’s tolerance to acute stress through electrical, thermal, and mechanical stimuli. The study also assessed circadian activity, climbing ability, and social behavior.

4. Synaptic Observation: Immunofluorescence was used to observe the morphology of neuromuscular junctions (NMJs) in Drosophila larvae to evaluate synaptic integrity.

5. Neurotransmitter Analysis: Ultra-high-performance liquid chromatography electrospray ionization quadrupole time-of-flight mass spectrometry (UHPLC-ESI-QTOF-MS) was employed to measure neurotransmitter levels.

Major Results and Supporting Data

1. Seizure-like Behavior and Acute Stress: The results showed that Drosophila with dYEATS2 knockdown (dYEATS2 ir-1 and dYEATS2 ir-2) exhibited significant seizure-like behaviors (SLB) under electrical, thermal, and mechanical stress, with recovery times significantly prolonged. This suggests that dYEATS2 plays an important role in regulating neuronal stress responses.

2. Complex Behavior Analysis: Drosophila with dYEATS2 knockdown showed defects in climbing ability and social behavior, though there were no significant changes in circadian activity. These findings suggest that dYEATS2 may play a role in neural plasticity and behavior regulation.

3. Neurotransmitter Levels: Mass spectrometry results demonstrated a significant reduction in dopamine (DA) levels in Drosophila with dYEATS2 knockdown, while levels of GABA, glutamate, and 5-HT remained unchanged. Further gene expression analysis indicated that dYEATS2 knockdown caused decreased expression of the gene encoding tyrosine hydroxylase (TH), a key enzyme in dopamine synthesis.

4. L-Dopa Rescue Experiment: Both chronic and acute administration of L-Dopa (a dopamine precursor) improved seizure-like behavior and locomotion in Drosophila with dYEATS2 knockdown. This further supports the hypothesis of dYEATS2’s role in maintaining dopaminergic synaptic integrity.

Conclusion and Research Significance

The findings indicate that dYEATS2 plays a crucial role in regulating dopamine synthesis and maintaining synaptic integrity. Its deficiency leads to neurological impairments and epileptic-like behavior in Drosophila. The rescuing effect of L-Dopa administration further supports this hypothesis. These discoveries not only enhance understanding of the pathogenesis of FAME4 but also provide new avenues for developing related therapeutic approaches.

Research Highlights

• Unveiling New Functions: This study is the first to reveal the role of dYEATS2 in regulating neuronal behavior and synaptic integrity.
• Dopamine Mechanism Analysis: The study elucidates in-depth the mechanism by which dYEATS2 regulates dopamine synthesis through TH gene expression.
• L-Dopa Rescue Experiments: Demonstrates that increasing dopamine levels can ameliorate associated neurological behavior defects.

Other Valuable Information

This research not only enhances the understanding of the function of the YEATS2 gene but also provides new perspectives for studying other related neurogenetic diseases. The use of the Drosophila model showcases the powerful potential of RNA interference technology in neurobehavioral research.

Review

Completed by an international and multidisciplinary team of experts, this study showcases high-level interdisciplinary collaboration. Through a series of rigorous experimental designs, the research reveals the essential role of the YEATS2 gene in the neural system of Drosophila. The results are scientifically valuable for understanding familial adult myoclonic epilepsy and developing potential treatments.