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.

journal_name

Brain Res

journal_title

Brain research

authors

Zhang C,Omran AG,He F,Deng X,Wu L,Peng J,Yin F

doi

10.1016/j.brainres.2013.10.053

subject

Has Abstract

pub_date

2014-01-16 00:00:00

pages

17-27

eissn

0006-8993

issn

1872-6240

pii

S0006-8993(13)01463-7

journal_volume

1543

pub_type

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