Arvaniti Maria, Jensen Majbrit M, Soni Neeraj, Wang Hong, Klein Anders B, Thiriet Nathalie, Pinborg Lars H, Muldoon Pretal P, Wienecke Jacob, Imad Damaj M, Kohlmeier Kristi A, Gondré-Lewis Marjorie C, Mikkelsen Jens D, Thomsen Morten S
Department of Drug Design & Pharmacology, University of Copenhagen, Copenhagen, Denmark.
Neurobiology Research Unit, University Hospital Copenhagen, Rigshospitalet, Copenhagen, Denmark.
J Neurochem. 2016 Sep;138(6):806-20. doi: 10.1111/jnc.13718. Epub 2016 Aug 15.
Nicotinic acetylcholine receptors (nAChRs) affect multiple physiological functions in the brain and their functions are modulated by regulatory proteins of the Lynx family. Here, we report for the first time a direct interaction of the Lynx protein LY6/PLAUR domain-containing 6 (Lypd6) with nAChRs in human brain extracts, identifying Lypd6 as a novel regulator of nAChR function. Using protein cross-linking and affinity purification from human temporal cortical extracts, we demonstrate that Lypd6 is a synaptically enriched membrane-bound protein that binds to multiple nAChR subtypes in the human brain. Additionally, soluble recombinant Lypd6 protein attenuates nicotine-induced hippocampal inward currents in rat brain slices and decreases nicotine-induced extracellular signal-regulated kinase phosphorylation in PC12 cells, suggesting that binding of Lypd6 is sufficient to inhibit nAChR-mediated intracellular signaling. We further show that perinatal nicotine exposure in rats (4 mg/kg/day through minipumps to dams from embryonic day 7 to post-natal day 21) significantly increases Lypd6 protein levels in the hippocampus in adulthood, which did not occur after exposure to nicotine in adulthood only. Our findings suggest that Lypd6 is a versatile inhibitor of cholinergic signaling in the brain, and that Lypd6 is dysregulated by nicotine exposure during early development. Regulatory proteins of the Lynx family modulate the function of nicotinic receptors (nAChRs). We report for the first time that the Lynx protein Lypd6 binds to nAChRs in human brain extracts, and that recombinant Lypd6 decreases nicotine-induced ERK phosphorylation and attenuates nicotine-induced hippocampal inward currents. Our findings suggest that Lypd6 is a versatile inhibitor of cholinergic signaling in the brain.
烟碱型乙酰胆碱受体（nAChRs）影响大脑中的多种生理功能，其功能受Lynx家族的调节蛋白调控。在此，我们首次报道了Lynx蛋白含LY6/PLAUR结构域6（Lypd6）与人脑提取物中的nAChRs直接相互作用，确定Lypd6为nAChR功能的新型调节因子。通过从人颞叶皮质提取物中进行蛋白质交联和亲和纯化，我们证明Lypd6是一种突触富集的膜结合蛋白，可与人脑中的多种nAChR亚型结合。此外，可溶性重组Lypd6蛋白可减弱大鼠脑片中尼古丁诱导的海马内向电流，并降低PC12细胞中尼古丁诱导的细胞外信号调节激酶磷酸化，这表明Lypd6的结合足以抑制nAChR介导的细胞内信号传导。我们进一步表明，大鼠围产期尼古丁暴露（从胚胎第7天到出生后第21天通过微型泵给母鼠注射4mg/kg/天）显著增加成年海马中Lypd6蛋白水平，而仅在成年期暴露于尼古丁后则不会出现这种情况。我们的研究结果表明，Lypd6是大脑中胆碱能信号的多功能抑制剂，并且Lypd6在早期发育过程中因尼古丁暴露而失调。Lynx家族的调节蛋白调节烟碱型受体（nAChRs）的功能。我们首次报道Lynx蛋白Lypd6与人脑提取物中的nAChRs结合，并且重组Lypd6可降低尼古丁诱导的ERK磷酸化并减弱尼古丁诱导的海马内向电流。我们的研究结果表明，Lypd6是大脑中胆碱能信号的多功能抑制剂。
