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.

Lynx1 and Aβ1–42 bind competitively to multiple nicotinic acetylcholine receptor subtypes

“…Here, we apply affinity purification with WsLynx1 to show that it forms complexes with nAChRs containing α3, α4, α5, α6, α7, β2, and β4 nAChR subunits in extracts from human cortex as well as rat cortex, midbrain and olfactory bulb. This is in stark contrast to the related Lynx protein, SLURP-1, which we have recently shown to selectively form complexes with the α7 subunit in human cortical extracts (Lyukmanova et al, 2016), and opens the possibility that Lynx1…”

Hereditary palmoplantar keratodermas. Part I. Non‐syndromic palmoplantar keratodermas: classification, clinical and genetic features

“…In 2001, Fisher et al . identified mutations in SLURP1 , encoding the secreted lymphocyte antigen 6 (LY6)/urokinase‐type plasminogen activator receptor (uPAR)‐related protein‐1 (Table ), in a diffuse PPK called mal de Meleda (MDM). MDM is an autosomal recessive PPK first described in patients from the Croatian island of Meleda (now called Mljet).…”

Cholinergic signalling mechanisms and early implant healing phases in healthy versus generalized aggressive periodontitis patients: A prospective, case–control study

“…Secreted Ly‐6/uPAR‐related protein 1 (SLURP‐1) is a member of the Ly6/uPAR family of proteins and shares some structural homology with nAChR agonists (Vasilyeva, Loktyushov, Bychkov, Shenkarev, & Lyukmanova, ). SLURP‐1 can act as allosteric antagonist of the α7nAChR but has also been demonstrated to activate metabotropic‐signalling pathways (Lyukmanova et al, ). SLURP‐1 is secreted at mucosal sites and has also been shown to have both immunomodulatory (Moriwaki et al, ) and antimicrobial properties (Moriwaki et al, ).…”

“…SLURP‐1 is secreted at mucosal sites and has also been shown to have both immunomodulatory (Moriwaki et al, ) and antimicrobial properties (Moriwaki et al, ). In addition, SLURP‐1 has been found to inhibit proliferation of oral keratinocytes (Lyukmanova et al, ). However, the exact roles of SLURP‐1 in oral infections have still to be delineated.…”