Loss of Quaking RNA binding protein disrupts the expression of genes associated with astrocyte maturation in mouse brain

Kristina Sakers,Eric Tycksen,Sean Brophy,Lorida Llaci,Michael J Vasek,Michael A Rieger,Yating Liu,Renate Lewis,Susan E Maloney,Joseph D Dougherty,Scott M Lee

doi:10.1038/s41467-021-21703-5

Abstract

Quaking RNA binding protein (QKI) is essential for oligodendrocyte development as myelination requires myelin basic protein mRNA regulation and localization by the cytoplasmic isoforms (e.g., QKI-6). QKI-6 is also highly expressed in astrocytes, which were recently demonstrated to have regulated mRNA localization. Here, we define the targets of QKI in the mouse brain via CLIPseq and we show that QKI-6 binds 3′UTRs of a subset of astrocytic mRNAs. Binding is also enriched near stop codons, mediated partially by QKI-binding motifs (QBMs), yet spreads to adjacent sequences. Using a viral approach for mosaic, astrocyte-specific gene mutation with simultaneous translating RNA sequencing (CRISPR-TRAPseq), we profile ribosome associated mRNA from QKI-null astrocytes in the mouse brain. This demonstrates a role for QKI in stabilizing CLIP-defined direct targets in astrocytes in vivo and further shows that QKI mutation disrupts the transcriptional changes for a discrete subset of genes associated with astrocyte maturation.

Highlights

Quaking RNA binding protein (QKI) is essential for oligodendrocyte development as myelination requires myelin basic protein mRNA regulation and localization by the cytoplasmic isoforms (e.g., QKI-6)
A few targets of QKI have been well studied in oligodendrocytes[18,20,24], and 6–10 mRNAs have been shown to be altered in cultured astrocytes and related tumors after siRNA knockdown[25,26], a compendium of all in vivo postnatal QKI targets has yet to be compiled
We chose to focus on QKI-6 for Cross-Linking and Immunoprecipitation (CLIP) experiments for two reasons: first, QKI5 is predominantly nuclear but QKI-6 and QKI-7 are both nuclear and cytoplasmic, and we posited that cytoplasmic isoforms are more likely candidates for translation regulation

Summary

Introduction

Quaking RNA binding protein (QKI) is essential for oligodendrocyte development as myelination requires myelin basic protein mRNA regulation and localization by the cytoplasmic isoforms (e.g., QKI-6). RBPs serve as the trans-acting factors that recognize cis-acting elements that are commonly found in the 3′UTR of localized transcripts[1] These proteins may directly transport RNAs to distal processes[2], alter transcript stability, or control translation[3,4] to prevent ectopic expression during mRNA transport or to activate translation in response to a molecular cue, for example. While these processes have been widely studied in other cell-types, it has recently become apparent that astrocytes must have substantial cytoplasmic post-transcriptional regulation: for example, a set of mRNAs enriched on ribosomes in peripheral astrocyte processes (PAPs) have been defined, consistent with sequence-regulated local translation[5]. In an astroglioma cell line, it has been shown that one QKI isoform stabilizes transcripts, those induced by interferons[23]

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Results

Conclusion