Ferric Chelate Reductase 1 Like Protein (FRRS1L) Associates with Dynein Vesicles and Regulates Glutamatergic Synaptic Transmission.

Wenyan Han,Shizhong Zhang,Huiqing Wang,Wei Lu,Jun Li

doi:10.3389/fnmol.2017.00402

Abstract

In the brain, AMPA receptors (AMPARs)-mediated excitatory synaptic transmission is critically regulated by the receptor auxiliary subunits. Recent proteomic studies have identified that Ferric Chelate Reductase 1 Like protein (FRRS1L), whose mutations in human lead to epilepsy, choreoathetosis, and cognitive deficits, is present in native AMPAR complexes in the brain. Here we have characterized FRRS1L in both heterologous cells and in mouse neurons. We found that FRRS1L interacts with both GluA1 and GluA2 subunits of AMPARs, but does not form dimers/oligomers, in HEK cells. In mouse hippocampal neurons, recombinant FRRS1L at the neuronal surface partially co-localizes with GluA1 and primarily localizes at non-synaptic membranes. In addition, native FRRS1L in hippocampus is localized at dynein, but not kinesin5B, vesicles. Functionally, over-expression of FRRS1L in hippocampal neurons does not change glutamatergic synaptic transmission. In contrast, single-cell knockout (KO) of FRRS1L strongly reduces the expression levels of the GluA1 subunit at the neuronal surface, and significantly decreases AMPAR-mediated synaptic transmission in mouse hippocampal pyramidal neurons. Taken together, these data characterize FRRS1L in heterologous cells and neurons, and reveal an important role of FRRS1L in the regulation of excitatory synaptic strength.

Highlights

In the brain, fast excitatory synaptic transmission is largely mediated by glutamate acting on a-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-type ionotropic glutamate receptors (AMPARs)
We found that HA-tagged Ferric Chelate Reductase 1 Like protein (FRRS1L) (HA-FRRS1L) could be co-immunoprecipitated with Flag-tagged GluA1 (Flag-GluA1) or Flag-tagged GluA2 (FlagGluA2) subunit of the AMPA-type ionotropic glutamate receptors (AMPARs) from cells co-transfected with both constructs, but not from control cells transfected with either one (Figure 1A), indicating that FRRS1L can associate with both GluA1 and GluA2 in human embryonic kidney 293T (HEK) cells
A substantial fraction of sHA-FRRS1L co-localized with a portion of endogenous surface GluA1 (Figure 1F), indicating that FRRS1L associates with a subpopulation of GluA1-containing AMPARs in hippocampal neurons

Summary

Introduction

Fast excitatory synaptic transmission is largely mediated by glutamate acting on AMPA-type ionotropic glutamate receptors (AMPARs). Accumulating studies over the last decade have shown that trafficking and function of AMPARs are critically controlled by an ever-growing list of membrane proteins, including TARPs (Chen et al, 2000; Tomita et al, 2003; Jackson and Nicoll, 2011), CNIHs (Schwenk et al, 2009), SynDig (Kalashnikova et al, 2010), CKAMPs/SHISAs (von Engelhardt et al, 2010; Farrow et al, 2015; Klaassen et al, 2016), GSG1L (Schwenk et al, 2012; Shanks et al, 2012; McGee et al, 2015; Gu et al, 2016a), PORCN (Erlenhardt et al, 2016), α/β-ABHD6 (Erlenhardt et al, 2016; Wei et al, 2016) and carnitine palmitoyltransferase 1C (CPT1C) (Gratacos-Batlle et al, 2014; Fado et al, 2015) These membrane proteins regulate AMPAR biogenesis, trafficking, gating, and pharmacological properties, and often play overlapping, and distinct, roles in the regulation of AMPAR-mediated synaptic transmission (Jackson and Nicoll, 2011; Straub and Tomita, 2012; Haering et al, 2014; Greger et al, 2017; Jacobi and von Engelhardt, 2017). It is important to understand the role of FRRS1L in the regulation of synaptic transmission

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