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ALS-linked mutations impair UBQLN2 stress-induced biomolecular condensate assembly in cells

View ORCID ProfileJulia F. Riley, View ORCID ProfileHeidi Hehnly, View ORCID ProfileCarlos A. Castañeda
doi: https://doi.org/10.1101/2020.10.17.335380
Julia F. Riley
1Department of Biology, Syracuse University, Syracuse, NY 13244
2Department of Chemistry, Syracuse University, Syracuse, NY 13244
3Interdisciplinary Neuroscience Program, Syracuse University, Syracuse, NY 13244
4Currently at the University of Pennsylvania, Neuroscience Graduate Group, Philadelphia, PA 19104
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Heidi Hehnly
1Department of Biology, Syracuse University, Syracuse, NY 13244
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Carlos A. Castañeda
1Department of Biology, Syracuse University, Syracuse, NY 13244
2Department of Chemistry, Syracuse University, Syracuse, NY 13244
3Interdisciplinary Neuroscience Program, Syracuse University, Syracuse, NY 13244
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  • For correspondence: cacastan@syr.edu
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Abstract

Mutations in Ubiquilin-2 (UBQLN2), a ubiquitin-binding shuttle protein involved in several protein quality control processes, can lead to amyotrophic lateral sclerosis (ALS). We previously found that wild-type UBQLN2 forms dynamic, membraneless biomolecular condensates upon cellular stress, and undergoes liquid-liquid phase separation in vitro. However, the impact of ALS-linked mutations on UBQLN2 condensate formation in cells is unknown. Here, we employ live-cell imaging with photokinetic analysis to investigate how five patient-derived ALS-linked mutations in UBQLN2 impact stress-induced UBQLN2 condensate assembly and condensate material properties. Both wild-type and mutant UBQLN2 condensates are generally cytoplasmic and liquid-like. However, cells transfected with mutant UBQLN2 contain fewer stress-induced UBQLN2 condensates than those with wild-type UBQLN2. Most strikingly, ectopically expressed P506T UBQLN2 forms the lowest number of stress-induced condensates of all UBQLN2 mutants, and these condensates are significantly smaller than those of wild-type UBQLN2. Fluorescence recovery after photobleaching (FRAP) analysis of UBQLN2 condensates revealed higher immobile fractions for UBQLN2 mutants, especially P506T. P497S and P497H mutations differentially impact condensate properties, demonstrating that the effects of ALS-linked mutations are both position- and amino acid-dependent. Collectively, our data show that disease mutations hinder assembly and alter viscoelastic properties of stress-induced UBQLN2 condensates, potentially leading to aggregates commonly observed in ALS.

Competing Interest Statement

The authors have declared no competing interest.

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The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
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Posted October 18, 2020.
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ALS-linked mutations impair UBQLN2 stress-induced biomolecular condensate assembly in cells
Julia F. Riley, Heidi Hehnly, Carlos A. Castañeda
bioRxiv 2020.10.17.335380; doi: https://doi.org/10.1101/2020.10.17.335380
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ALS-linked mutations impair UBQLN2 stress-induced biomolecular condensate assembly in cells
Julia F. Riley, Heidi Hehnly, Carlos A. Castañeda
bioRxiv 2020.10.17.335380; doi: https://doi.org/10.1101/2020.10.17.335380

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