# KCC2 phosphorylation dynamics relate to oligomerization during development and after spinal cord injury

## TL;DR

This study explores how phosphorylation of KCC2 affects its oligomerization during development and after spinal cord injury in rats.

## Contribution

The study identifies tyrosine dephosphorylation at the membrane as a key correlate of KCC2 oligomerization.

## Key findings

- KCC2 moves from the cytoplasm to the membrane during development with increased intracellular phosphorylation.
- Spinal cord injury shifts KCC2 phosphorylation toward lower oligomers and higher membrane tyrosine phosphorylation.
- DOI treatment partially reverses these effects, promoting oligomerization and developmental-like phosphorylation.

## Abstract

Effective synaptic inhibition relies on KCC2, a K+-Cl− cotransporter that forms oligomers to extrude chloride. However, how phosphorylation regulates oligomerization remains unclear. To address this, we studied the phosphorylation of KCC2 in the lumbar spinal cord of neonatal rats by subcellular fractionation and phospho-specific western blotting across three paradigms with distinct expression profiles: developmental upregulation, spinal cord injury (SCI)-induced downregulation, and pharmacological rescue with the 5-HT2A/2C agonist DOI. KCC2 moved from cytoplasm to membrane during development, exhibiting increased intracellular phosphorylation at serine, threonine, and tyrosine residues and decreased membrane tyrosine phosphorylation as a result of the oligomer formation. SCI caused a shift in this ratio in favor of lower oligomers, higher membrane tyrosine phosphorylation and lower intracellular threonine phosphorylation. DOI partially reversed these deficits, reinforcing oligomerization and adapting phosphorylation to a developmental profile. In sum, this study identifies tyrosine dephosphorylation at the membrane as a reliable correlate of oligomerization and offers a unified quantitative model for the regulation of KCC2 in development, after injury, and upon pharmacological rescue.