ABSTRACT
SFPQ is a nuclear RNA-binding protein that is involved in a wide range of physiological processes including neuronal development and homeostasis. However, the mislocalization and cytoplasmic aggregation of SFPQ are associated with the pathophysiology of amyotrophic lateral sclerosis (ALS). We have previously reported that zinc mediates SFPQ polymerization and promotes the formation of cytoplasmic aggregates in neurons. Here we characterize two familial ALS (fALS)-associated SFPQ variants, which cause amino acid substitutions in the proximity of the SFPQ zinc-coordinating center (N533H and L534I). Both mutants display increased zinc-binding affinities, which can be explained by the presence of a secondary zinc-binding site revealed by the 1.83Å crystal structure of the human SFPQ L534I mutant. Overexpression of these fALS-associated mutants significantly increases the number of SFPQ cytoplasmic aggregates in primary neurons. Although they do not affect the density of dendritic spines, the presence of SFPQ cytoplasmic aggregates causes a marked reduction in the levels of the GluA1, but not the GluA2 subunit of AMPA-type glutamate receptors on the neuronal surface. Taken together, our data demonstrate that fALS-associated mutations enhance the propensity of SFPQ to bind zinc and form aggregates, leading to the dysregulation of AMPA receptor subunit composition, which may contribute to neuronal dysfunction in ALS.
Competing Interest Statement
The authors have declared no competing interest.
Footnotes
Funding This study was supported by grants from the Motor Neurone Disease Research Australia (the Judy Mitchell MND Research Grant) to VA, ML and JW, and the Australian Medical Research Future Fund (Clem Jones Centre for Ageing Dementia Research Flagship Project Grant) to VA. ML holds a Tracey Banivanua Mar Fellowship (La Trobe University). JW holds a University of Queensland (UQ) Amplify Fellowship. JZAT was supported by a UQ Researcher Retention (RSA2) Fellowship. JH was the recipient of a PhD scholarship from La Trobe University.
Conflict of interest The authors declare no conflict of interests.
Ethical Approval All animal handling procedures were carried out in accordance with the Australian Code of Practice for the Care and Use of Animals for Scientific Purposes and were approved by the University of Queensland Animal Ethics Committee (AEC approval number QBI/047/18).
Informed Consent This article does not contain any studies with human participants performed by any of the authors. All authors have read and agreed with the submission of the manuscript.
Data Availability Atomic coordinates and structure factors for the reported crystal structures have been deposited with the Protein Data Bank under the accession number 7SP0.
Abbreviations
- ALS
- amyotrophic lateral sclerosis
- AMPA
- α-amino-3-hydroxy-5-methyl-4- isoxazoleproprionic acid
- AMPAR
- AMPA receptor
- DBHS
- Drosophila behavior human splicing
- fALS
- familial ALS
- FUS
- fused in sarcoma
- RBP
- RNA-binding protein
- r.m.s.d.
- root mean square deviation
- RRM
- RNA-recognition motif
- RXRα
- retinoic X receptor α
- SFPQ
- splicing factor proline- and glutamine-rich
- TDP-43
- trans-activation response element DNA-binding protein 43
- WT
- wild-type.