Neuronal cell depolarization induces intragenic chromatin modifications affecting NCAM alternative splicing

Area of Science:

• Neuroscience
• Molecular Biology
• Epigenetics

Background:

• Alternative splicing is crucial for neuronal diversity and function.
• The kinetic coupling between transcription and splicing influences alternative splicing outcomes.
• Understanding how external stimuli regulate splicing in neurons is essential.

Purpose of the Study:

• To investigate the physiological pathways linking membrane depolarization to alternative splicing of the neural cell adhesion molecule (NCAM).
• To elucidate the role of chromatin modifications in regulating NCAM alternative splicing during neuronal activity.

Main Methods:

• Neuronal cell culture and membrane depolarization.
• Analysis of NCAM mRNA splicing patterns using RT-PCR.
• Chromatin immunoprecipitation (ChIP) to assess histone modifications (H3K9 acetylation, H3K36 tri-methylation).
• Treatment with histone deacetylase inhibitors (trichostatin A).

Main Results:

• Membrane depolarization induces skipping of exon 18 in NCAM mRNA, independent of CaMKIV.
• Depolarization leads to specific H3K9 hyper-acetylation within the NCAM gene, associated with chromatin relaxation and H3K36 tri-methylation.
• Histone modifications and splicing changes are reversible upon withdrawal of depolarization.
• Histone deacetylase inhibition mimics and potentiates depolarization-induced splicing changes.

Conclusions:

• Neuronal membrane depolarization regulates NCAM alternative splicing through transcription-splicing coupling.
• Intragenic epigenetic modifications, specifically histone acetylation and methylation, mediate this response.
• These findings reveal a mechanism linking neuronal activity to gene expression regulation relevant for neuronal differentiation and function.