RESUMO
TWIK1 belongs to a family of K(+) channels involved in neuronal excitability and cell volume regulation. Its tissue distribution suggests a role in epithelial potassium transport. Here we show that TWIK1 is expressed in a subapical compartment in renal proximal tubules and in polarized MDCK cells. In nonpolarized cells, this compartment corresponds to pericentriolar recycling endosomes. We identified EFA6, an exchange factor for the small G protein ADP-ribosylation factor 6 (ARF6), as a protein binding to TWIK1. EFA6 interacts with TWIK1 only when it is bound to ARF6. Because ARF6 modulates endocytosis at the apical surface of epithelial cells, the ARF6/EFA6/TWIK1 association is probably important for channel internalization and recycling.
Assuntos
Fatores de Ribosilação do ADP/metabolismo , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Fatores de Alongamento de Peptídeos/metabolismo , Canais de Potássio/metabolismo , Fator 6 de Ribosilação do ADP , Animais , Endocitose/fisiologia , Imunofluorescência , Guanosina Difosfato/metabolismo , Guanosina Trifosfato/metabolismo , Células HeLa , Humanos , Rim/metabolismo , Camundongos , Proteínas do Tecido Nervoso , Transferrina/metabolismoRESUMO
The perception of photic information and its translation into a rhythmic melatonin signal differ considerably among vertebrates. In the fish pineal gland, melatonin biosynthesis is controlled directly by the natural light/dark cycle. There are indications that the mechanisms of phototransduction are similar in the retinal and pineal photoreceptor cells. Here we report the molecular cloning of a novel ionic cyclic guanosine monophosphate (cGMP)-gated channel from trout pineal photoreceptors. The deduced amino acid sequence exhibits a high sequence homology to cyclic nucleotide-gated-3 (CNG) channels from retinal cones. In situ hybridization with sections of trout pineal gland revealed the expression of CNG channel in photoreceptor cells of the pineal organ. Electrophysiological studies by means of patch-clamp technique indicated that the native channel in photoreceptor cells and the expressed channel in a human cell line (HEK 293 cells) have properties similar to those of cone-CNG (cCNG)-3 channels. They are activated by cGMP, insensitive to cyclic adenosine monophosphate (cAMP) and blocked by intracellular Mg2+ ions at positive voltage values. They have a single-channel conductance close to 42 pS in negative voltage range. In transfected HEK cells loaded with the calcium indicator dye Fura 2, direct activation of CNG channels by 8-Br-cGMP increased fluorescence. The signal was blocked by the addition of Mg2+ ions. From these results, it is suggested that the pineal cyclic nucleotide-gated channel is a good candidate for mediating calcium entry into the pineal photoreceptors. It is most probably a key element in the signalling pathways that control the rhythmic production of melatonin.
