title: The cholinergic immune regulation mediated by a novel muscarinic acetylcholine receptor through TNF pathway in oyster Crassostrea gigas.
abstract: Muscarinic receptors, which selectively take muscarine as their ligand, are critical for the immunological and physiological processes in animals. In the present study, the open region frame (ORF) of a homologue of muscarinic acetylcholine (ACh) receptor (mAChR) was amplified from oyster Crassostrea gigas (named as CgmAChR-1), whose full length was 1983 bp and the protein it encoded contained 660 amino acids with a seven transmembrane region. Phylogeny analysis suggested that CgmAChR-1 shared homology with M5 muscarinic receptor found in invertebrates including Habropoda laboriosa, Acromyrmex echinatior and Echinococcus granulosus. After cell transfection of CgmAChR-1 into HEK293T cells and ACh incubation, the level of intracellular Ca(2+) and cAMP increased significantly (p < 0.05). Such trend could be reverted with the addition of M3 and M5 muscarinic receptor antagonists DAMP and DAR. The CgmAChR-1 transcripts were ubiquitously detectable in seven different tissues with the maximal expression level in adductor muscle. When the oysters received LPS stimulation, CgmAChR-1 mRNA expression in haemocyte was increased to the highest level (6.05-fold, p < 0.05) at 24 h, while blocking CgmAChR-1 using receptor antagonists before LPS stimulation promoted the expression of oyster TNF, resulting in the increase of haemocyte apoptosis index. These results suggested that CgmAChR-1 was the key molecule in cholinergic neuroendocrine-immune system contributing to the regulation of TNF expression and apoptosis process.
authors.affiliation: Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, Qingdao, 266071, China; University of Chinese Academy of Sciences, Beijing, 100049, China.
authors.forename: Zhaoqun
authors.initials: Z
authors.lastname: Liu
authors.affiliation: Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, Qingdao, 266071, China.
authors.forename: Zhi
authors.initials: Z
authors.lastname: Zhou
authors.affiliation: Key Laboratory of Mariculture & Stock Enhancement in North China's Sea, Ministry of Agriculture, Dalian Ocean University, Dalian, 116023, China. Electronic address: wanglingling@ms.qdio.ac.cn.
authors.forename: Lingling
authors.initials: L
authors.lastname: Wang
authors.affiliation: Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, Qingdao, 266071, China.
authors.forename: Wenjing
authors.initials: W
authors.lastname: Dong
authors.affiliation: Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, Qingdao, 266071, China.
authors.forename: Limei
authors.initials: L
authors.lastname: Qiu
authors.affiliation: Key Laboratory of Mariculture & Stock Enhancement in North China's Sea, Ministry of Agriculture, Dalian Ocean University, Dalian, 116023, China. Electronic address: lshsong@dlou.edu.cn.
authors.forename: Linsheng
authors.initials: L
authors.lastname: Song
mesh_terms: D000818:Animals; D017209:Apoptosis; D002118:Calcium / Q000378:metabolism; D018678:Cholinergic Agents / Q000378:metabolism; D003001:Cloning, Molecular; D049896:Crassostrea / Q000276:immunology*; D000242:Cyclic AMP / Q000378:metabolism; D057809:HEK293 Cells; D006434:Hemocytes / Q000502:physiology*; D006801:Humans; D009116:Muscarine / Q000378:metabolism; D011976:Receptors, Muscarinic / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
chemical_list: D018678:Cholinergic Agents; D011976:Receptors, Muscarinic; D014409:Tumor Necrosis Factor-alpha; D009116:Muscarine; D000242:Cyclic AMP; D002118:Calcium
keywords: Acetylcholine; Crassostrea gigas; Immune regulation; Muscarinic acetylcholine receptor; TNF pathway