Human Three-Finger Protein Lypd6 Is a Negative Modulator of the Cholinergic System in the Brain

Lypd6 is a GPI-tethered protein from the Ly-6/uPAR family expressed in the brain. Lypd6 enhances the Wnt/beta-catenin signaling, although its action on nicotinic acetylcholine receptors (nAChRs) have been also proposed. To investigate a cholinergic activity of Lypd6, we studied a recombinant water-soluble variant of the human protein (ws-Lypd6) containing isolated three-finger LU-domain. Experiments at different nAChR subtypes expressed in Xenopus oocytes revealed the negative allosteric modulatory activity of ws-Lypd6. Ws-Lypd6 inhibited ACh-evoked currents at alpha 3 beta 4- and alpha 7-nAChRs with IC50 of similar to 35 and 10 mu M, respectively, and the maximal amplitude of inhibition of 30-50%. EC50 of ACh at alpha 3 beta 4-nAChRs (similar to 30 mu M) was not changed in the presence of 35 mu M ws-Lypd6, while the maximal amplitude of ACh-evoked current was reduced by similar to 20%. Ws-Lypd6 did not elicit currents through nAChRs in the absence of ACh. Application of 1 mu M ws-Lypd6 significantly inhibited (up to similar to 28%) choline-evoked current at alpha 7-nAChRs in rat hippocampal slices. Similar to snake neurotoxin alpha-bungarotoxin, ws-Lypd6 suppressed the long-term potentiation (LTP) in mouse hippocampal slices. Colocalization of endogenous GPI-tethered Lypd6 with alpha 3 beta 4- and alpha 7-nAChRs was detected in primary cortical and hippocampal neurons. Ws-Lypd6 interaction with the extracellular domain of alpha 7-nAChR was modeled using the ensemble protein-protein docking protocol. The interaction of all three Lypd6 loops (fingers) with the entrance to the orthosteric ligand-binding site and the loop C of the primary receptor subunit was predicted. The results obtained allow us to consider Lypd6 as the endogenous negative modulator involved in the regulation of the cholinergic system in the brain.

Automated summary

Lypd6 is a GPI-tethered protein in the Ly-6/uPAR family that enhances Wnt/beta-catenin signaling and has been suggested to affect nicotinic acetylcholine receptors (nAChRs). The recombinant water-soluble variant of Lypd6 inhibits ACh-evoked currents at α3β4- and α7-nAChRs, and can also suppress long-term potentiation in mouse hippocampal slices. Colocalization of Lypd6 with nAChRs was observed in primary cortical and hippocampal neurons, indicating its role in cholinergic system regulation in the brain.