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

  • Front Cell Dev Biol. 2021 Sep 21;9:662227. doi: 10.3389/fcell.2021.662227.
Dmitrii Kulbatskii  1 ,  Zakhar Shenkarev  2  3 ,  Maxim Bychkov  1 ,  Eugene Loktyushov  1 ,  Mikhail Shulepko  1 ,  Sergey Koshelev  4 ,  Igor Povarov  5 ,  Alexander Popov  4  6 ,  Steve Peigneur  7 ,  Anton Chugunov  2  3  8 ,  Sergey Kozlov  4 ,  Irina Sharonova  5 ,  Roman Efremov  2  3  8 ,  Vladimir Skrebitsky  5 ,  Jan Tytgat  7 ,  Mikhail Kirpichnikov  1  9 ,  Ekaterina Lyukmanova  1  3  9
Affiliations
  • 1. Bioengineering Department, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry Russian Academy of Sciences (RAS), Moscow, Russia.
  • 2. Structural Biology Department, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry Russian Academy of Sciences (RAS), Moscow, Russia.
  • 3. Phystech School of Biological and Medical Physics, Moscow Institute of Physics and Technology, Moscow, Russia.
  • 4. Department of Molecular Neurobiology, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry Russian Academy of Sciences (RAS), Moscow, Russia.
  • 5. Brain Research Department, Research Center of Neurology, Moscow, Russia.
  • 6. Institute of Neuroscience, Nizhny Novgorod University, Nizhny Novgorod, Russia.
  • 7. Toxicology and Pharmacology, University of Leuven (KU Leuven), Leuven, Belgium.
  • 8. International Laboratory for Supercomputer Atomistic Modelling and Multi-Scale Analysis, National Research University Higher School of Economics, Moscow, Russia.
  • 9. Biological Faculty, Lomonosov Moscow State University, Moscow, Russia.
PMID: 34631692 DOI: 10.3389/fcell.2021.662227
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.

Keywords
Ly6/uPAR; Lynx1; Lypd6; Lypd6b; cognitive function; nAChR; synaptic plasticity; three-finger.