Interaction of Synaptosomal-Associated Protein 25 with Neutral Sphingomyelinase 2: Functional Impact on the Sphingomyelin Pathway. (10th February 2020)

• Record Type: Journal Article
• Authors: Won, Jong Hoon; Jeon, Hyung Jun; Kim, Seok Kyun; Shin, In Chul; Jang, Ji Min; Ha, Hae Chan; Back, Moon Jung; Kim, Dae Kyong
• Is Part Of: Neuroscience. Volume 427(2020)
• Page Start: 1
• Page End: 15
• Publication Date: 2020-02-10
• Subjects: BSA bovine serum albumin -- co-IP co-immunoprecipitation -- DAPI 4′, 6-diamidino-2-phenylindole -- DEAE diethylaminoethanol -- EDTA ethylenediaminetetraacetic acid -- HPLC high-performance liquid chromatography -- IgG immunoglobulin G -- IP immunoprecipitation -- LC/MS/MS liquid chromatography and tandem mass spectrometry -- nSMase neutral sphingomyelinase -- PBS phosphate-buffered saline -- PLA proximity ligation assay -- PVDF polyvinylidene difluoride -- RPMI Roswell Park Memorial Institute -- SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis -- siRNA small interfering RNA -- SMase sphingomyelinase -- SNAP-25 synaptosomal-associated protein 25 -- SNARE soluble N-ethylmaleimide-sensitive factor attachment protein receptor; synaptosomal-associated protein 25 -- ceramide -- neutral sphingomyelinase 2 -- dopamine -- neurotransmission; Neurochemistry -- Periodicals; Neurophysiology -- Periodicals; Neurology -- Periodicals; Neurochimie -- Périodiques; Neurophysiologie -- Périodiques; Neurochemistry; Neurophysiology; Electronic journals; Periodicals; Electronic journals; 612.8
• DOI: 10.1016/j.neuroscience.2019.08.015
• Languages: English
• ISSNs: 0306-4522

Abstract

Highlights: We examined protein-protein interaction between SNAP-25 and nSMase2 in neuron. SNAP-25 mediates translocation of nSMase2 and regulates ceramide formation. Dopamine release is mediated by ceramide production. Abstract: Neurotransmitter release is mediated by ceramide, which is generated by sphingomyelin hydrolysis. In the present study, we examined whether synaptosomal-associated protein 25 (SNAP-25) is involved in ceramide production and exocytosis. Neutral sphingomyelinase 2 (nSMase2) was partially purified from bovine brain and we found that SNAP-25 was enriched in the nSMase2-containing fractions. In rat synaptosomes and PC12 cells, the immunoprecipitation pellet of anti-SNAP-25 antibody showed higher nSMase activity than the immunoprecipitation pellet of anti-nSMase2 antibody. In PC12 cells, SNAP-25 was colocalized with nSMase2. Transfection of SNAP-25 small interfering RNA (siRNA) significantly inhibited nSMase2 translocation to the plasma membrane. A23187-induced ceramide production was concomitantly reduced in SNAP-25 siRNA-transfected PC12 cells compared with that in scrambled siRNA-transfected cells. Moreover, transfection of SNAP-25 siRNA inhibited dopamine release, whereas addition of C6 -ceramide to the siRNA-treated cells moderately reversed this inhibition. Additionally, nSMase2 inhibition reduced dopamine release. Collectively, our results indicate that SNAP-25 interacts with nSMase2 during ceramide production, which mediates exocytosis and neurotransmitter release.