Abstract
Lypd6 is a GPI-tethered protein from the Ly-6/uPAR family expressed in the brain. Lypd6 enhances the Wnt/β-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 α3β4- and α7-nAChRs with IC50 of ∼35 and 10 μM, respectively, and the maximal amplitude of inhibition of 30–50%. EC50 of ACh at α3β4-nAChRs (∼30 μM) was not changed in the presence of 35 μM ws-Lypd6, while the maximal amplitude of ACh-evoked current was reduced by ∼20%. Ws-Lypd6 did not elicit currents through nAChRs in the absence of ACh. Application of 1 μM ws-Lypd6 significantly inhibited (up to ∼28%) choline-evoked current at α7-nAChRs in rat hippocampal slices. Similar to snake neurotoxin α-bungarotoxin, ws-Lypd6 suppressed the long-term potentiation (LTP) in mouse hippocampal slices. Colocalization of endogenous GPI-tethered Lypd6 with α3β4- and α7-nAChRs was detected in primary cortical and hippocampal neurons. Ws-Lypd6 interaction with the extracellular domain of α7-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.
Highlights
Nicotinic acetylcholine receptors are ligand-gated ion channels and important participants of signaling in the nervous, endocrine, and immune systems of mammals (Wessler and Kirkpatrick, 2009)
The auxiliary regulatory proteins have been described for the AMPA, NMDA, GABAA receptors, and nicotinic acetylcholine receptors (nAChRs)
The GPI-anchored Ly-6/urokinase-type plasminogen activator (uPAR) protein Lynx1 is colocalized with α7-nAChRs in the brain areas important for learning and memory (Ibañez-Tallon et al, 2002), and is crucial for the loss of neuronal plasticity during postnatal development (Morishita et al, 2010) and regulation of the spine turnover in the adult visual cortex (Sajo et al, 2016)
Summary
Nicotinic acetylcholine receptors (nAChRs) are ligand-gated ion channels and important participants of signaling in the nervous, endocrine, and immune systems of mammals (Wessler and Kirkpatrick, 2009). The most known members of the Ly-6/uPAR family are snake α-neurotoxins and the receptor of urokinase-type plasminogen activator (uPAR), containing one and three LU-domains, respectively (Vasilyeva et al, 2017). There are a lot of regulatory Ly-6/uPAR proteins containing the single LU-domain, and some of them are responsible for modulation of nAChRs in mammals (Ibañez-Tallon et al, 2002; Tekinay et al, 2009; Jensen et al, 2015; Arvaniti et al, 2016; Lyukmanova et al, 2016a,b). For some of the Ly-6/uPAR proteins (e.g., Lynx, PCSA, CD59, uPAR), both soluble and membrane-tethered forms have been reported (Fletcher et al, 1994; Thomsen et al, 2014; Jensen et al, 2015; Su et al, 2016)
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