ABSTRACT
Loss of the RNA binding protein FMRP causes Fragile X Syndrome (FXS), the most common cause of inherited intellectual disability, yet it is unknown how FMRP function varies across brain regions and cell types and how this contributes to disease pathophysiology. Here we use conditional tagging of FMRP and CLIP (FMRP cTag CLIP) to examine FMRP mRNA targets in hippocampal CA1 pyramidal neurons, a critical cell type for learning and memory relevant to FXS phenotypes. Integrating these data with analysis of ribosome-bound transcripts in these neurons revealed CA1-enriched binding of autism-relevant mRNAs, and CA1-specific regulation of transcripts encoding circadian proteins. This contrasted with different targets in cerebellar granule neurons, and was consistent with circadian defects in hippocampus-dependent memory in Fmr1 knockout mice. These findings demonstrate differential FMRP-dependent regulation of mRNAs across neuronal cell types that may contribute to phenotypes such as memory defects and sleep disturbance associated with FXS.
Subject(s)
Autistic Disorder/metabolism , CA1 Region, Hippocampal/metabolism , Fragile X Mental Retardation Protein/genetics , Fragile X Syndrome/genetics , Memory Disorders/genetics , Pyramidal Cells/metabolism , Animals , Autistic Disorder/genetics , Autistic Disorder/physiopathology , CA1 Region, Hippocampal/cytology , Cerebellum/cytology , Cerebellum/metabolism , Circadian Clocks/genetics , Circadian Clocks/physiology , Disease Models, Animal , Fragile X Mental Retardation Protein/metabolism , Fragile X Syndrome/metabolism , Fragile X Syndrome/physiopathology , Gene Expression Regulation , Humans , Memory Disorders/metabolism , Memory Disorders/physiopathology , Mice, Inbred C57BL , Mice, Knockout , Neurons/metabolismABSTRACT
RNA localization to neuronal dendrites and axons is increasingly recognized as a significant and widespread mechanism of gene expression control in neurons. High-throughput RNA sequencing is rapidly expanding the universe of known localized mRNAs. Although there are inherent difficulties in preparing sequencing libraries from dendrites and axons in the context of intact brain, genetic labeling strategies have paved the way for improved studies of this type. As the list of localized mRNAs grows, there is increasing need for functional validation of localized transcripts-that is, do particular localized transcripts serve demonstrable physiologic functions in axons or dendrites? Finally, specific details about what localized mRNAs do once they reach distal processes have long been elusive. Recent work using single-molecule imaging and other techniques is starting to fill in the picture of how transcripts navigate the localized environment and undergo activity-dependent translational de-repression. © 2017 Wiley Periodicals, Inc. Develop Neurobiol 78: 331-339, 2018.
Subject(s)
Neurons/metabolism , RNA Transport/physiology , AnimalsABSTRACT
FMRP loss of function causes Fragile X syndrome (FXS) and autistic features. FMRP is a polyribosome-associated neuronal RNA-binding protein, suggesting that it plays a key role in regulating neuronal translation, but there has been little consensus regarding either its RNA targets or mechanism of action. Here, we use high-throughput sequencing of RNAs isolated by crosslinking immunoprecipitation (HITS-CLIP) to identify FMRP interactions with mouse brain polyribosomal mRNAs. FMRP interacts with the coding region of transcripts encoding pre- and postsynaptic proteins and transcripts implicated in autism spectrum disorders (ASD). We developed a brain polyribosome-programmed translation system, revealing that FMRP reversibly stalls ribosomes specifically on its target mRNAs. Our results suggest that loss of a translational brake on the synthesis of a subset of synaptic proteins contributes to FXS. In addition, they provide insight into the molecular basis of the cognitive and allied defects in FXS and ASD and suggest multiple targets for clinical intervention.
