Abstract
Purpose :
Olfactomedin 1 (Olfm1) is a component of the AMPA receptor complex in the vertebrate brain and retina. Olfm1a and olfm1b double knockout zebrafish (olfm1null) demonstrated changes in the retinal structure and visual motor function (ARVO 2014, Abstract #1918670). Changes in the composition of the AMPA receptor complex in the olfm1 null fish were investigated to elucidate the molecular mechanisms of Olfm1 action in the brain and retina.
Methods :
Brains of adult fish were used to isolate synaptosomal and lipid raft fractions. Olfm1, GluR2 and SNARE complex proteins were immunoprecipitated with specific antibodies and analyzed by Western blotting. GluR2 internalization was investigated in retinal sections of adult fish.
Results :
In the adult wild-type (wt) brain, Olfm1 was preferentially localized to the synaptosomal membrane fraction together with GluR2, PSD95, Syntaxin and SNAP25. In this fraction, Olfm1 interacted with Syntaxin, SNAP25, Synaptophysin, VAMP2 and GluR2, as judged by co-immunoprecipitation indicating participation of Olfm1 in both pre- and post-synaptic events. The palmitoylation of GluR2 was greatly reduced in the synaptosomal fraction of olfm1 null fish compared with wt. Olfm1 co-precipitated with GluR2 was also palmitoylated. Changes in palmitoylation may cause reduced internalization of GluR2, as observed in retinal explant cultures from olfm1a/b null larvae compared with wt. Analysis of the composition of lipid raft prepared from adult brain demonstrated the presence of Olfm1. The levels of several proteins (GluR2, SNAP25, Flotillin1, VAMP2) were significantly reduced in the raft fraction isolated from olfm1null brain compared with wt.
Conclusions :
Zebrafish Olfm1 is a synaptic protein that interacts with the GluR2 and SNARE complexes. Olfm1 may be involved in the regulation of trafficking of some synaptic proteins and their localization to the lipid raft in the brain and retina.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.