Trafficking properties and activity regulation of the neuronal glycine transporter GLYT2 by protein kinase C

Amparo Fornés , Enrique Núñez , Pablo Alonso-Torres , Carmen Aragón , Beatriz López-Corcuera (1)

1 Biología Molecular

The neuronal glycine transporter GLYT2 controls the availability of the neurotransmitter in glycinergic synapses and the modulation of its function may influence synaptic transmission. Active transporter is located in membrane rafts and reaches the cell surface through intracellular trafficking. Here we prove that GLYT2 constitutively recycles between the cell interior and the plasma membrane by means of a monensin-sensitive trafficking pathway. Besides, a regulated trafficking can be triggered by 4α-phorbol 12-myristate 13-acetate (PMA). We demonstrate that PMA inhibits GLYT2 transport by causing net accumulation of the protein in internal compartments through an increase of the internalization rate. In addition, a small increase of plasma membrane delivery and a redistribution of the transporter to non-raft domains is triggered by PMA. A previously identified phorbol ester-resistant mutant (K422E) displaying an acidic substitution in a regulatory site, exhibits constitutive traffic but, in contrast to wild type, fails to show glycine uptake inhibition, membrane raft redistribution and trafficking modulation by PMA. We prove that the action of PMA on GLYT2 involves protein kinase C (PKC)-dependent and independent pathways, although an important fraction of the effects are PKC–mediated. We show the additional participation of signaling pathways triggered by the small GTPase Rac1 on PMA action. GLYT2 inhibition by PMA and monensin take also place in brainstem primary neurons and synaptosomes, pointing to a GLYT2 trafficking regulation in the central nervous system.

Mots clés: Life Sciences

Amparo Fornés, Enrique Núñez, Pablo Alonso-Torres, Carmen Aragón, Beatriz López-Corcuera. Trafficking properties and activity regulation of the neuronal glycine transporter GLYT2 by protein kinase C. Biochemical Journal, 2008, 412 (3), pp.495-506. ⟨10.1042/BJ20071018⟩. ⟨hal-00478857⟩