title: Human Secreted Ly-6/uPAR Related Protein-1 (SLURP-1) Is a Selective Allosteric Antagonist of α7 Nicotinic Acetylcholine Receptor.
abstract: SLURP-1 is a secreted toxin-like Ly-6/uPAR protein found in epithelium, sensory neurons and immune cells. Point mutations in the slurp-1 gene cause the autosomal inflammation skin disease Mal de Meleda. SLURP-1 is considered an autocrine/paracrine hormone that regulates growth and differentiation of keratinocytes and controls inflammation and malignant cell transformation. The majority of previous studies of SLURP-1 have been made using fusion constructs containing, in addition to the native protein, extra polypeptide sequences. Here we describe the activity and pharmacological profile of a recombinant analogue of human SLURP-1 (rSLURP-1) differing from the native protein only by one additional N-terminal Met residue. rSLURP-1 significantly inhibited proliferation (up to ~ 40%, EC50 ~ 4 nM) of human oral keratinocytes (Het-1A cells). Application of mecamylamine and atropine,--non-selective inhibitors of nicotinic acetylcholine receptors (nAChRs) and muscarinic acetylcholine receptors, respectively, and anti-α7-nAChRs antibodies revealed α7 type nAChRs as an rSLURP-1 target in keratinocytes. Using affinity purification from human cortical extracts, we confirmed that rSLURP-1 binds selectively to the α7-nAChRs. Exposure of Xenopus oocytes expressing α7-nAChRs to rSLURP-1 caused a significant non-competitive inhibition of the response to acetylcholine (up to ~ 70%, IC50 ~ 1 μM). It was shown that rSLURP-1 binds to α7-nAChRs overexpressed in GH4Cl cells, but does not compete with 125I-α-bungarotoxin for binding to the receptor. These findings imply an allosteric antagonist-like mode of SLURP-1 interaction with α7-nAChRs outside the classical ligand-binding site. Contrary to rSLURP-1, other inhibitors of α7-nAChRs (mecamylamine, α-bungarotoxin and Lynx1) did not suppress the proliferation of keratinocytes. Moreover, the co-application of α-bungarotoxin with rSLURP-1 did not influence antiproliferative activity of the latter. This supports the hypothesis that the antiproliferative activity of SLURP-1 is related to 'metabotropic' signaling pathway through α7-nAChR, that activates intracellular signaling cascades without opening the receptor channel.
authors.affiliation: Biological Department, Lomonosov Moscow State University, Moscow, Russian Federation.|Department of Bioengineering, Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russian Federation.
authors.forename: Ekaterina N
authors.initials: EN
authors.lastname: Lyukmanova
authors.affiliation: Biological Department, Lomonosov Moscow State University, Moscow, Russian Federation.|Department of Bioengineering, Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russian Federation.
authors.forename: Mikhail A
authors.initials: MA
authors.lastname: Shulepko
authors.affiliation: Department of Molecular Basics of Neurosignalling, Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russian Federation.
authors.forename: Denis
authors.initials: D
authors.lastname: Kudryavtsev
authors.affiliation: Biological Department, Lomonosov Moscow State University, Moscow, Russian Federation.|Department of Bioengineering, Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russian Federation.
authors.forename: Maxim L
authors.initials: ML
authors.lastname: Bychkov
authors.affiliation: Biological Department, Lomonosov Moscow State University, Moscow, Russian Federation.|Department of Bioengineering, Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russian Federation.
authors.forename: Dmitrii S
authors.initials: DS
authors.lastname: Kulbatskii
authors.affiliation: Department of Molecular Basics of Neurosignalling, Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russian Federation.
authors.forename: Igor E
authors.initials: IE
authors.lastname: Kasheverov
authors.affiliation: Department of Structural Biology, Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russian Federation.
authors.forename: Maria V
authors.initials: MV
authors.lastname: Astapova
authors.affiliation: Biological Department, Lomonosov Moscow State University, Moscow, Russian Federation.|Department of Structural Biology, Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russian Federation.
authors.forename: Alexey V
authors.initials: AV
authors.lastname: Feofanov
authors.affiliation: Department of Drug Design and Pharmacology, University of Copenhagen, Copenhagen, Denmark.|Neurobiology Research Unit, University Hospital, Copenhagen, Copenhagen, Denmark.
authors.forename: Morten S
authors.initials: MS
authors.lastname: Thomsen
authors.affiliation: Neurobiology Research Unit, University Hospital, Copenhagen, Copenhagen, Denmark.
authors.forename: Jens D
authors.initials: JD
authors.lastname: Mikkelsen
authors.affiliation: Biological Department, Lomonosov Moscow State University, Moscow, Russian Federation.|Department of Structural Biology, Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russian Federation.|Moscow Institute of Physics and Technology (State University), Dolgoprudny, Moscow Region, Russian Federation.
authors.forename: Zakhar O
authors.initials: ZO
authors.lastname: Shenkarev
authors.affiliation: Department of Molecular Basics of Neurosignalling, Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russian Federation.
authors.forename: Victor I
authors.initials: VI
authors.lastname: Tsetlin
authors.affiliation: Biological Department, Lomonosov Moscow State University, Moscow, Russian Federation.|Department of Bioengineering, Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russian Federation.
authors.forename: Dmitry A
authors.initials: DA
authors.lastname: Dolgikh
authors.affiliation: Biological Department, Lomonosov Moscow State University, Moscow, Russian Federation.|Department of Bioengineering, Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russian Federation.
authors.forename: Mikhail P
authors.initials: MP
authors.lastname: Kirpichnikov
mesh_terms: D000494:Allosteric Regulation / Q000187:drug effects / Q000235:genetics; D000950:Antigens, Ly / Q000235:genetics / Q000494:pharmacology; D002460:Cell Line; D006801:Humans; D015603:Keratinocytes / Q000166:cytology / Q000378:metabolism*; D011485:Protein Binding; D011994:Recombinant Proteins / Q000235:genetics / Q000494:pharmacology; D014568:Urokinase-Type Plasminogen Activator / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D064569:alpha7 Nicotinic Acetylcholine Receptor / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism*
chemical_list: D000950:Antigens, Ly; D011994:Recombinant Proteins; C048963:SLURP1 protein, human; D064569:alpha7 Nicotinic Acetylcholine Receptor; D014568:Urokinase-Type Plasminogen Activator