Developmentally regulated KCC2 phosphorylation is essential for dynamic GABA-mediated inhibition and survival

This is the author accepted manuscript. The final version is available from American Association for the Advancement of Science via the DOI in this recordData and materials availability: All data needed to evaluate the conclusions in the paper are present in the paper or the Supplementary Materials.Despite its importance for γ-aminobutyric acid (GABA) inhibition and involvement in neurodevelopmental disease, the regulatory mechanisms of the K+/Cl− cotransporter KCC2 (encoded by SLC12A5) during maturation of the central nervous system (CNS) are not entirely understood. Here, we applied quantitative phosphoproteomics to systematically map sites of KCC2 phosphorylation during CNS development in the mouse. KCC2 phosphorylation at Thr906 and Thr1007, which inhibits KCC2 activity, underwent dephosphorylation in parallel with the GABA excitatory-inhibitory sequence in vivo. Knockin mice expressing the homozygous phosphomimetic KCC2 mutations T906E/T1007E (Kcc2E/E), which prevented the normal developmentally regulated dephosphorylation of these sites, exhibited early postnatal death from respiratory arrest and a marked absence of cervical spinal neuron respiratory discharges. Kcc2E/E mice also displayed disrupted lumbar spinal neuron locomotor rhythmogenesis and touch-evoked status epilepticus associated with markedly impaired KCC2-dependent Cl− extrusion. These data identify a previously unknown phosphorylation-dependent KCC2 regulatory mechanism during CNS development that is essential for dynamic GABA-mediated inhibition and survival.March of DimesSimons FoundationNIHMinistry of Education, Culture, Sports, Science and Technology of JapanJapan Society for the Promotion of Scienc