title: The N-terminal HSDCIF motif is required for cell surface trafficking and dimerization of family B G protein coupled receptor PAC1.
abstract: PAC1 is PACAP (pituitary adenylate cyclase-activating polypeptide) preferring receptor belonging to class B G protein coupled receptor (GPCR) mediating the most effects of PACAP. The important role of G protein coupled receptor homo/heteromerization in receptor folding, maturation, trafficking, and cell surface expression has become increasingly evident. The bimolecular fluorescence complementation (BiFC) and bioluminescence resonance energy transfer (BRET) assay were used in this research to confirm the dimerization of PAC1 for the first time. The structure-activity relationship focused on the N-terminal HSDCIF motif, which locates behind the signal sequence and has high homology with PACAP (1-6), was assayed using a receptor mutant with the deletion of the HSDCIF motif. The fluorescence confocal microscope observation showed that the deletion of the HSDCIF motif impaired the cell delivery of PAC1. The results of BiFC, BRET and westernblot indicated that the deletion of HSDCIF motif and the replacement of the Cys residue with Ala in HSDCIF motif resulted in the disruption of receptor dimerization. And the exogenous chemically synthesized oligopeptide HSDCIF (100 nmol/L) not only down-regulated the dimerization of PAC1, induced the internalization of PAC1, but also inhibited the proliferation of CHO cells expressing PAC1 stably and decreased the activity of PACAP on the cell viability. All these data suggested that the N-terminal HSDCIF motif played key role in the trafficking and the dimerization of PAC1, and the exogenous oligopeptide HSDCIF had effects on the cell signaling, trafficking and the dimerization of PAC1.
authors.affiliation: Cell Biology Institute, the Department of Cell Biology, Jinan University, Guangzhou, China. rongjie_yu1123@163.com
authors.forename: Rongjie
authors.initials: R
authors.lastname: Yu
authors.forename: Xiaoling
authors.initials: X
authors.lastname: Guo
authors.forename: Jiaping
authors.initials: J
authors.lastname: Zhong
authors.forename: Mei
authors.initials: M
authors.lastname: Li
authors.forename: Zhixing
authors.initials: Z
authors.lastname: Zeng
authors.forename: Huahua
authors.initials: H
authors.lastname: Zhang
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D060807:Bioluminescence Resonance Energy Transfer Techniques; D015153:Blotting, Western; D016466:CHO Cells; D049109:Cell Proliferation; D006224:Cricetinae; D003412:Cricetulus; D005109:Extracellular Matrix; D005455:Fluorescent Antibody Technique; D006801:Humans; D008969:Molecular Sequence Data; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D051237:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide, Type I / Q000737:chemistry* / Q000378:metabolism*; D015398:Signal Transduction; D013329:Structure-Activity Relationship
chemical_list: D051237:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide, Type I