Cell-specific exon methylation and CTCF binding in neurons regulates calcium ion channel splicing and function

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Abstract

Cell-specific alternative splicing modulates myriad cell functions and is disrupted in disease. The mechanisms governing alternative splicing are known for relatively few genes and typically focus on RNA splicing factors. In sensory neurons, cell-specific alternative splicing of the presynaptic Ca_V channel Cacna1b gene modulates opioid sensitivity. How this splicing is regulated is unknown. We find that cell and exon -specific DNA hypomethylation permits CTCF binding, the master regulator of mammalian chromatin structure, which, in turn, controls splicing in a DRG-derived cell line. In vivo, hypomethylation of an alternative exon specifically in nociceptors, likely permits CTCF binding and expression of Ca_V2.2 channel isoforms with increased opioid sensitivity in mice. Following nerve injury, exon methylation is increased, and splicing is disrupted. Our studies define the molecular mechanisms of cell-specific alternative splicing of a functionally validated exon in normal and disease states – and reveal a potential target for the treatment of chronic pain.

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All data generated or analysed during this study are included in the manuscript and supporting files.

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Eduardo Javier López Soto

Neuroscience, Brown University, Providence, United States

Competing interests
The authors declare that no competing interests exist.
Diane Lipscombe

Neuroscience, Brown University, Providence, United States

For correspondence
diane_lipscombe@brown.edu

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-7146-9119

Funding

National Institute of Neurological Disorders and Stroke (NS055251)

Diane Lipscombe

Warren Alpert Foundation

Eduardo Javier López Soto

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Ethics

Animal experimentation: Mice were housed and bred at Brown University. All protocols and procedures were approved by the Brown University Institutional Animal Care and Use Committee (IACUC # 1706000275).Mice were anesthetized with 3% isoflurane, and all effort was made to minimize suffering.

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This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.