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The membrane trafficking and functionality of the K + ‐Cl − co‐transporter KCC2 is regulated by TGF‐β2
Author(s) -
Khakipoor Shokoufeh,
Roussa Eleni
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.1007.55
Subject(s) - microbiology and biotechnology , creb , hippocampal formation , chemistry , gene knockdown , signal transduction , biology , neuroscience , transcription factor , biochemistry , gene
GABA A response shifts from depolarizing to hyperpolarizing during neuronal maturation and it is well established that K + ‐Cl − ‐cotransporter KCC2 up‐regulation is responsible for this process. Failure in KCC2 up‐regulation is seen in epilepsy and stroke. Surprisingly, there is not much known about regulatory pathways involved in the KCC2 expression, trafficking, and functional expression. Here, we investigate the effect of transforming growth factor beta 2, TGF‐β2, in the regulation of KCC2 trafficking and functional expression in mouse mature hippocampal neurons and analyze the underlying molecular pathways. Therefore embryonic day 18 primary hippocampal neurons and hindbrain slices were used. KCC2 protein and surface expression is investigated by immunoblotting, surface biotinylation and immunofluorescence. Signaling pathways are studied using siRNA knockdown, co‐immunoprecipitation, and 3D STED microscopy. KCC2 functional expression is measured by somatodendritic Cl − gradient recording via whole cell patch clamp. The results show that KCC2 surface and functional expression is significantly increased in hippocampal neurons treated with TGF‐β2, compared to non‐treated cells. TGF‐β2 treatment leads to cAMP response element‐binding protein (CREB) phosphorylation and Ras‐associated binding protein 11b (Rab11b) gene expression and also increases Rab11b/KCC2 interaction and co‐localization. Knocking down either CREB or Rab11b suppresses KCC2 membrane and functional expression. Pre‐Bötzinger complex neurons in TGF‐β2 deficient mice have a significant decrease in KCC2 membrane expression, compared to wildtype and exogenous TGFβ2 application can rescue this phenotype. We introduce TGF‐β2/CREB/Rab11b as a novel regulatory pathway involved in KCC2 trafficking and functional activity in mouse mature hippocampal neurons. Support or Funding Information This work was supported by grants from the Deutsche Forschungsgemeinschaft [grant numbers KR1477/15‐1, SFB 780 TP B02 to K.K. and E.R.].