A cytoplasmic quaking I isoform regulates the hnRNP F/H-dependent alternative splicing pathway in myelinating glia.

M D Mandler,Y Feng,L Ku

doi:10.1093/nar/gku353

Abstract

The selective RNA-binding protein quaking I (QKI) plays important roles in controlling alternative splicing (AS). Three QKI isoforms are broadly expressed, which display distinct nuclear-cytoplasmic distribution. However, molecular mechanisms by which QKI isoforms control AS, especially in distinct cell types, still remain elusive. The quakingviable (qkv) mutant mice carry deficiencies of all QKI isoforms in oligodendrocytes (OLs) and Schwann cells (SWCs), the myelinating glia of central and peripheral nervous system (CNS and PNS), respectively, resulting in severe dysregulation of AS. We found that the cytoplasmic isoform QKI-6 regulates AS of polyguanine (G-run)-containing transcripts in OLs and rescues aberrant AS in the qkv mutant by repressing expression of two canonical splicing factors, heterologous nuclear ribonucleoproteins (hnRNPs) F and H. Moreover, we identified a broad spectrum of in vivo functional hnRNP F/H targets in OLs that contain conserved exons flanked by G-runs, many of which are dysregulated in the qkv mutant. Interestingly, AS targets of the QKI-6-hnRNP F/H pathway in OLs are differentially affected in SWCs, suggesting that additional cell-type-specific factors modulate AS during CNS and PNS myelination. Together, our studies provide the first evidence that cytoplasmic QKI-6 acts upstream of hnRNP F/H, which forms a novel pathway to control AS in myelinating glia.

Highlights

Alternative splicing (AS) allows production of multiple mRNAs from a single gene that often encode distinct protein isoforms to perform paradoxically opposite functions [1]
Our previous studies showed that quaking I (QKI)-6 is necessary for normal AS of the proteolipid protein (PLP) pre-mRNA expressed in OLs, which is a well-characterized target of heterologous nuclear ribonucleoproteins (hnRNPs) F/H [12,25,34]
To test whether QKI-6 may interact with hnRNP F/H mRNAs in myelinating OLs in vivo, we performed UV-crosslinking immunoprecipitation (CLIP) to isolate mRNA complexes formed with cytoplasmic QKI-6 from the brain of transgenic mice that express FLAG-QKI6 in OLs [23]

Summary

Introduction

Alternative splicing (AS) allows production of multiple mRNAs from a single gene that often encode distinct protein isoforms to perform paradoxically opposite functions [1]. The majority of splicing factors are ubiquitously expressed [1], yet alternatively spliced mRNA isoforms are often differentially regulated during development and in different cell types [5,6]. Dysregulation of AS in OLs causes human neurological diseases, as seen in the aberrant AS of proteolipid protein (PLP) pre-mRNA in a familial form of Pelizaeus–Merzbacher disease [9]. Severe dysregulation of AS is caused by deficiency of the quaking I (QKI) RNA-binding protein in myelinating glia of the homozygous quakingviable (qkv/qkv) mutant mouse, represented by the pre-mRNAs of PLP and myelin associated glycoprotein (MAG) [10,11,12]

Methods

Results

Conclusion