FMRP Control of Ribosome Translocation Promotes Chromatin Modifications and Alternative Splicing of Neuronal Genes Linked to Autism

Abstract

<jats:title>Summary</jats:title><jats:p>Silencing of <jats:italic>FMR1</jats:italic> and loss of its gene product FMRP results in Fragile X Syndrome. FMRP binds brain mRNAs and inhibits polypeptide elongation. Using ribosome profiling of the hippocampus, we find that ribosome footprint levels in <jats:italic>Fmr1</jats:italic>-deficient tissue mostly reflect changes in RNA abundance. Profiling over a time course of ribosome runoff in wildtype tissue reveals a wide range of ribosome translocation rates; on many mRNAs, the ribosomes are stalled. Sucrose gradient ultracentrifugation of hippocampal slices after ribosome runoff reveals that FMRP co-sediments with stalled ribosomes; and its loss results in decline of ribosome stalling on specific mRNAs. One such mRNA encodes SETD2, a lysine methyltransferase that catalyzes H3K36me3. ChIP-Seq demonstrates that loss of FMRP alters the deployment of this epigenetic mark on chromatin. H3K36me3 is associated with alternative pre-RNA processing, which we find occurs in an FMRP-dependent manner on transcripts linked to neural function and autism spectrum disorders.</jats:p><jats:sec><jats:title>Highlights</jats:title><jats:p><jats:list list-type="simple"><jats:list-item><jats:p>- Loss of FMRP results in decline of ribosome stalling on specific mRNAs (eg., SETD2)</jats:p></jats:list-item><jats:list-item><jats:p>- Increased SETD2 protein levels alter H3K36me3 marks in FMRP deficient hippocampus</jats:p></jats:list-item><jats:list-item><jats:p>- Genome-wide changes in mRNA alternative splicing occur in FMRP deficient hippocampus</jats:p></jats:list-item><jats:list-item><jats:p>- H3K36me3 marks and alternative splicing changes occur on transcripts linked to autism</jats:p></jats:list-item></jats:list></jats:p></jats:sec>

Disome and Trisome Profiling Reveal Genome-wide Targets of Ribosome Quality Control

“…Indeed, recent studies have utilized disome profiling in yeast, mouse, and human cells (Arpat et al, 2019;Han et al, 2020;Tuck et al, 2020;Zhao et al, 2019). In addition, ribosome stalling has been shown to be regulated by the RNA-binding protein FMRP, which is known to be critical for neuronal development (Darnell et al, 2011;Das Sharma et al, 2019;Shah et al, 2020). Future disome profiling experiments can therefore be used to explore how dysregulated disome formation leads to disease phenotypes.…”

The molecular biology of FMRP: new insights into fragile X syndrome

“…However, TE does not allow for the assessment of mRNAs associated with stalled ribosomes (that is, ribosomes that have initiated translation but that become stalled during the process of polypeptide elongation) (see below). Several studies have used ribosome profiling to investigate translational control by FMRP in various tissues and cells 36,58,59,60,61,62,63 . A case in point is adult mouse neural stem cells (aNSCs); they give rise to neurons and glia but when FMRP is absent, the cells over-proliferate and primarily differentiate into more glia at the expense of neurons 64,65 .…”

Do Fragile X Syndrome and Other Intellectual Disorders Converge at Aberrant Pre-mRNA Splicing?

“…In FMRP deficient hippocampus tissues, one of the mRNAs released from translational repression is SETD2, resulting in increase of SETD2 (a lysine methyltransferase) protein levels. Increased SETD2 alters the H3k36me3 marks on the chromatin, which correlated with alternative splicing defects in mRNAs critical for proper synaptic function (5).…”

“…(4)]. Detailed molecular analysis of targets of translational de-repression in FXS led to the finding that a chromatin modifier, SETD2 (SET domain containing methyltransferase protein), is altered, resulting in downstream alterations in the chromatin landscape and genome-wide aberrant alternative splicing of mRNAs (5). Aberrant alternative splicing has previously been studied in complex NDDs such as ASD (6)(7)(8) and other psychiatric illnesses with a neurodevelopmental trajectory including Schizophrenia (8,9), Bipolar Disorder (8, 9), Huntington's (10), and recently in monogenic IDs such as FXS (5), PTEN (11), and RTT (12-14) (Table 1).…”