title: Screening and identification of dynamin-1 interacting proteins in rat brain synaptosomes.
abstract: Dynamin-1 is a multi-domain GTPase that is crucial for the fission stage of synaptic vesicle recycling and vesicle trafficking. In this study, we constructed prokaryotic expression plasmids for the four functional domains of dynamin-1, which are pGEX-4T-2-PH, pGEX-4T-2-PRD, pGEX-4T-2-GED and pGEX-4T-2-GTPase. Glutathione S-transferase pull-down, co-immunoprecipitation (co-IP), and liquid chromatography/mass spectrometry were used to screen and identify dynamin-1 interacting proteins in rat brain synaptosomes. We identified a set of 63 candidate protein interactions, including 36 proteins interacting with dynamin-1 C-terminal proline-rich domain (PRD), 14 with pleckstrin-homology domain (PH), 7 with GTPase effector domain (GED) and 6 with GTPase domain, consisting of synaptic vesicle-associated proteins, cytoskeletal proteins, metabolic enzymes and other proteins. We selected three previously unreported dynamin-1 interacting proteins to verify their interaction with dynamin-1 under native conditions. Using co-IP, we found that Rab GDP-dissociation inhibitor (Rab GDI) and chloride channel 3 (ClC-3) do interact with dynamin-1, but not with TUC-4b (the TOAD-64/Ulip/CRMP (TUC) family member). Those novel interactions detected in our study offer valuable insight into the protein-protein interacting network that could enhance our understanding of dynamin-1 mediated synaptic vesicle recycling.
authors.affiliation: Department of Pediatrics, Xiangya Hospital, Central South University, 87 Xiangya Road, Hunan 410008, PR China. Electronic address: zhangciliu@163.com.
authors.forename: Ciliu
authors.initials: C
authors.lastname: Zhang
authors.affiliation: Department of Pediatrics, Xiangya Hospital, Central South University, 87 Xiangya Road, Hunan 410008, PR China. Electronic address: agomran1@yahoo.com.
authors.forename: Ahmed Galal
authors.initials: AG
authors.lastname: Omran
authors.affiliation: Department of Pediatrics, Xiangya Hospital, Central South University, 87 Xiangya Road, Hunan 410008, PR China. Electronic address: bubbly_ho@163.com.
authors.forename: Fang
authors.initials: F
authors.lastname: He
authors.affiliation: Department of Pediatrics, Xiangya Hospital, Central South University, 87 Xiangya Road, Hunan 410008, PR China. Electronic address: dengxiaolu1019@yahoo.cn.
authors.forename: Xiaolu
authors.initials: X
authors.lastname: Deng
authors.affiliation: Department of Pediatrics, Xiangya Hospital, Central South University, 87 Xiangya Road, Hunan 410008, PR China. Electronic address: wulei8509@hotmail.com.
authors.forename: Lei
authors.initials: L
authors.lastname: Wu
authors.affiliation: Department of Pediatrics, Xiangya Hospital, Central South University, 87 Xiangya Road, Hunan 410008, PR China. Electronic address: pengjing4346@gmail.com.
authors.forename: Jing
authors.initials: J
authors.lastname: Peng
authors.affiliation: Department of Pediatrics, Xiangya Hospital, Central South University, 87 Xiangya Road, Hunan 410008, PR China. Electronic address: yf2323@hotmail.com.
authors.forename: Fei
authors.initials: F
authors.lastname: Yin
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000818:Animals; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D001921:Brain / Q000648:ultrastructure*; D018118:Chloride Channels / Q000378:metabolism; D062126:Databases, Chemical; D034284:Dynamin I / Q000378:metabolism*; D020690:GTPase-Activating Proteins / Q000378:metabolism; D020730:Guanine Nucleotide Dissociation Inhibitors / Q000378:metabolism; D047468:Immunoprecipitation; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000378:metabolism; D060066:Protein Interaction Maps; D051381:Rats; D013574:Synaptosomes / Q000378:metabolism*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; D018118:Chloride Channels; C115376:ClC-3 channel; C086796:GDP dissociation inhibitor 1; D020690:GTPase-Activating Proteins; D020730:Guanine Nucleotide Dissociation Inhibitors; D008869:Microtubule-Associated Proteins; D009419:Nerve Tissue Proteins; C568776:Phlda1 protein, rat; C075637:amphiphysin; D034284:Dynamin I
keywords: Dynamin-1; Interacting protein; Rat; Synaptosome