title: The membrane trafficking and functionality of the K+-Cl- co-transporter KCC2 is regulated by TGF-β2.
abstract: Functional activation of the neuronal K(+)-Cl(-) co-transporter KCC2 (also known as SLC12A5) is a prerequisite for shifting GABAA responses from depolarizing to hyperpolarizing during development. Here, we introduce transforming growth factor β2 (TGF-β2) as a new regulator of KCC2 membrane trafficking and functional activation. TGF-β2 controls membrane trafficking, surface expression and activity of KCC2 in developing and mature mouse primary hippocampal neurons, as determined by immunoblotting, immunofluorescence, biotinylation of surface proteins and KCC2-mediated Cl(-) extrusion. We also identify the signaling pathway from TGF-β2 to cAMP-response-element-binding protein (CREB) and Ras-associated binding protein 11b (Rab11b) as the underlying mechanism for TGF-β2-mediated KCC2 trafficking and functional activation. TGF-β2 increases colocalization and interaction of KCC2 with Rab11b, as determined by 3D stimulated emission depletion (STED) microscopy and co-immunoprecipitation, respectively, induces CREB phosphorylation, and enhances Rab11b gene expression. Loss of function of either CREB1 or Rab11b suppressed TGF-β2-dependent KCC2 trafficking, surface expression and functionality. Thus, TGF-β2 is a new regulatory factor for KCC2 functional activation and membrane trafficking, and a putative indispensable molecular determinant for the developmental shift of GABAergic transmission.
authors: Institute of Anatomy and Cell Biology, Department of Molecular Embryology, Faculty of Medicine, University of Freiburg, Albertstrasse 17, Freiburg D-79104, Germany Institute of Anatomy and Cell Biology, Department of Neuroanatomy, Faculty of Medicine, University of Freiburg, Albertstrasse 17, Freiburg D-79104, Germany kerstin.krieglstein@anat.uni-freiburg.de eleni.roussa@anat.uni-freiburg.de.
authors: Eleni
authors: 0000-0002-0495-1597
authors: E
authors: Roussa
authors: Institute of Anatomy and Cell Biology, Department of Molecular Embryology, Faculty of Medicine, University of Freiburg, Albertstrasse 17, Freiburg D-79104, Germany.
authors: Jan Manuel
authors: JM
authors: Speer
authors: Institute of Anatomy and Cell Biology, Department of Molecular Embryology, Faculty of Medicine, University of Freiburg, Albertstrasse 17, Freiburg D-79104, Germany.
authors: Ilona
authors: I
authors: Chudotvorova
authors: Institute of Anatomy and Cell Biology, Department of Molecular Embryology, Faculty of Medicine, University of Freiburg, Albertstrasse 17, Freiburg D-79104, Germany.
authors: Shokoufeh
authors: S
authors: Khakipoor
authors: Institute of Biotechnology, University of Helsinki, Viikinkaari 9, Helsinki FIN-00014, Finland.
authors: Sergei
authors: 0000-0001-5475-4156
authors: S
authors: Smirnov
authors: Institute of Biotechnology, University of Helsinki, Viikinkaari 9, Helsinki FIN-00014, Finland.
authors: Claudio
authors: C
authors: Rivera
authors: Institute of Anatomy and Cell Biology, Department of Molecular Embryology, Faculty of Medicine, University of Freiburg, Albertstrasse 17, Freiburg D-79104, Germany kerstin.krieglstein@anat.uni-freiburg.de eleni.roussa@anat.uni-freiburg.de.
authors: Kerstin
authors: K
authors: Krieglstein
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000378:metabolism*; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D005260:Female; D006624:Hippocampus / Q000166:cytology; D006801:Humans; D042541:Intracellular Space / Q000378:metabolism; D008297:Male; D008810:Mice, Inbred C57BL; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D027981:Symporters / Q000187:drug effects / Q000378:metabolism*; D053781:Transforming Growth Factor beta2 / Q000494:pharmacology*; D020691:rab GTP-Binding Proteins / Q000378:metabolism; D000096922:K Cl- Cotransporters; D000099193:rab11 GTP-Binding Proteins
chemical_list: D017362:Cyclic AMP Response Element-Binding Protein; D027981:Symporters; D053781:Transforming Growth Factor beta2; D020691:rab GTP-Binding Proteins; D000096922:K Cl- Cotransporters; D000099193:rab11 GTP-Binding Proteins
keywords: CREB; Growth factor; KCC2; Neuronal development; Rab11b