Activation of group-I metabotropic glutamate receptors impairs stress granules formation and enhances cellular damage under stress condition.

Fragile X Syndrome (FXS), a common form of inherited intellectual disability and autism, is caused by the lack of Fragile X Mental Retardation Protein (FMRP), an RNA binding protein involved in RNA metabolism and protein synthesis. Upon cellular stress, FMRP is associated with the pool of mRNAs that are recruited and protected into stress granules (SGs), ribonucleoproteic structures containing translation components, like eIF2?, and RNA binding proteins, including FMRP. During stress, canonical protein synthesis is blocked and FMRP may transport mRNAs into SGs. Group-I metabotropic glutamate receptors (mGluRs) are implicated in the pathophysiology of FXS; their activation stimulate FMRP-mediated RNA transport and protein synthesis, but their role in SGs formation has never been investigated. To this aim, wild type (WT) and Fmr1 knockout (KO) cultured astrocytes were treated with the group-I-mGluR agonist DHPG (100 uM) for 5 minutes and then exposed to sodium arsenite (NaAsO2, 200-500 µM) for 60-90 minutes. We used: immunocytochemistry for TIA-1 protein, a marker of SGs, and FMRP to detect SGs and FMRP recruitment in SGs; Western blot to evaluate phosphorylation of eIF2? and FMRP, two crucial key events in SG formation and modulation of protein synthesis, respectively; MTT colorimetric assay to assess the effect of DHPG on stress-induced damage. We observed a lower number of cells exhibiting SGs in Fmr1 KO than WT astrocytes after exposure to NaAsO2. In WT cultures the activation of group-I mGluRs before stress reduced SGs formation, TIA-1/FMRP co-localization in SGs, phosphorylation of eIF2? and FMRP, whilst increased stress-induced cytotoxicity. In contrast, group-I GluR activation had no effect in KO astrocytes. In conclusion, FMRP plays a key role in SGs formation and mGluR activation affects SGs formation through a FMRP-mediated mechanism. Phosphorylation of eIF2? initiates SGs formation and phosphorylated-FMRP is crucial for FMRP-mediated inhibition of protein synthesis; thus, mGluRs may act by shifting the balance from inhibition to activation of protein synthesis during stress. This mechanism may also be involved in group-I mGlu receptor sensitization to oxidative-stress induced damage.