FMRP phosphorylation modulates neuronal translation through YTHDF1

Translation activation of local synaptic mRNA is critical to learning and memory. Despite extensive studies on how phosphorylations of ribosome units and translation factors enable fast response to exogenous stimuli, our knowledge on molecular pathways utilized by RNA binding proteins (RBPs) to control translation of specific mRNAs remains incomplete. We have previously found that YTHDF1 regulates depolarization-induced protein synthesis by promoting translation of N6-methyladenosine (m6A)-modified transcripts. Here we report an unexpected mechanism that the stimuli-induced neuronal translation is mediated by phosphorylation of a YTHDF1-binding protein FMRP. Phosphorylation of FMRP serine 499 induced by neuronal depolarization alters the condensing behavior of prion-like protein YTHDF1. Unphosphorylated FMRP sequesters YTHDF1 away from the translation initiation complex, whereas the stimulation-induced FMRP phosphorylation releases YTHDF1 to form translational active condensates with the ribosome to activate translation of YTHDF1 target transcripts. In fragile X syndrome (FXS) models characterized with low FMRP expression, we observed YTHDF1-mediated hyperactive translation, which notably impacts FXS pathophysiology. Developmental defects in an FXS forebrain organoid model could be reversed by a selective small-molecule inhibitor of YTHDF1 which acts by suppressing its condensation in neurons. We characterized transcriptome-wide translation alterations with the inhibitor treatment in organoids and identified targets that explain alleviated FXS pathology. Our study thus reveals FMRP and its phosphorylation as an important regulator of the activity-dependent translation during neuronal development and stimulation, and identifies YTHDF1 as a potential therapeutic target for FXS in which developmental defects caused by FMRP depletion could be reversed through YTHDF1 inhibition. [Dataset 1] 16 samples. Duplicates for mRNA-Seq and riboLace ribosome-protected fragments samples in isolated primary neurons from P1-3 wildtype, Fmr1 knockout mouse treated by YTHDF1 inhibitor SAC or DMSO control [Dataset 2] 16 samples. Duplicates for mRNA-Seq and riboLace ribosome-protected fragments samples in cultured control or FXS human forebrain organoids treated by SAC or DMSO