TIA1 variant drives myodegeneration in multisystem proteinopathy with SQSTM1 mutations
YouJin Lee, … , Conrad C. Weihl, Bjarne Udd
YouJin Lee, … , Conrad C. Weihl, Bjarne Udd
Published February 19, 2018
Citation Information: J Clin Invest. 2018;128(3):1164-1177. https://doi.org/10.1172/JCI97103.
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Research Article Genetics Muscle biology

TIA1 variant drives myodegeneration in multisystem proteinopathy with SQSTM1 mutations

Abstract

Multisystem proteinopathy (MSP) involves disturbances of stress granule (SG) dynamics and autophagic protein degradation that underlie the pathogenesis of a spectrum of degenerative diseases that affect muscle, brain, and bone. Specifically, identical mutations in the autophagic adaptor SQSTM1 can cause varied penetrance of 4 distinct phenotypes: amyotrophic lateral sclerosis (ALS), frontotemporal dementia, Paget’s disease of the bone, and distal myopathy. It has been hypothesized that clinical pleiotropy relates to additional genetic determinants, but thus far, evidence has been lacking. Here, we provide evidence that a TIA1 (p.N357S) variant dictates a myodegenerative phenotype when inherited, along with a pathogenic SQSTM1 mutation. Experimentally, the TIA1-N357S variant significantly enhances liquid-liquid–phase separation in vitro and impairs SG dynamics in living cells. Depletion of SQSTM1 or the introduction of a mutant version of SQSTM1 similarly impairs SG dynamics. TIA1-N357S–persistent SGs have increased association with SQSTM1, accumulation of ubiquitin conjugates, and additional aggregated proteins. Synergistic expression of the TIA1-N357S variant and a SQSTM1-A390X mutation in myoblasts leads to impaired SG clearance and myotoxicity relative to control myoblasts. These findings demonstrate a pathogenic connection between SG homeostasis and ubiquitin-mediated autophagic degradation that drives the penetrance of an MSP phenotype.

Authors

YouJin Lee, Per Harald Jonson, Jaakko Sarparanta, Johanna Palmio, Mohona Sarkar, Anna Vihola, Anni Evilä, Tiina Suominen, Sini Penttilä, Marco Savarese, Mridul Johari, Marie-Christine Minot, David Hilton-Jones, Paul Maddison, Patrick Chinnery, Jens Reimann, Cornelia Kornblum, Torsten Kraya, Stephan Zierz, Carolyn Sue, Hans Goebel, Asim Azfer, Stuart H. Ralston, Peter Hackman, Robert C. Bucelli, J. Paul Taylor, Conrad C. Weihl, Bjarne Udd

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Figure 6

SQSTM1 disease mutations alter SG kinetics and synergistically mediate myotoxicity with TIA1-N357S.

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SQSTM1 disease mutations alter SG kinetics and synergistically mediate m...
(A) Bar graph showing the percentage of MEFs containing endogenous TIA1-positive SGs. MEFs were transfected with mCherry, mCherry-SQSTM1-WT, or 1 of 3 different disease mutations (PL, MV, or AX) incubated at 42°C for 1 hour and subsequently returned to 37°C for the indicated durations. Transfected cells were counted and are indicated as the total number of cells. Representative data were pooled from 3 independent experiments (n = 450~550). (B) Bar graph showing the percentage of fibroblasts containing TIA1/G3BP1-positive SGs from fibroblasts of control patients (fibroblast lines 112 and 409), patients carrying the TIA1-N357S variant (fibroblast lines 107 and 319), and a patient carrying both a SQSTM1-A390X mutation and a TIA1-N357S variant (fibroblast line 483) immediately following 0.5 mM AsIII treatment for 1 hour or following a 40-minute recovery. (C) Immunofluorescence images of patients’ fibroblasts detailed in B, immunostained with TIA1 (green) and G3BP1 antibodies (red) before, immediately following 0.5 mM AsIII treatment for 1 hour, and following a 40-minute recovery. DAPI nuclear staining is shown in blue. Scale bars: 5 μm. Representative data were pooled from 3 independent experiments (n = 120~150). (D) Bar graph showing the percentage of C2C12 myoblasts containing TIA1-positive SGs. C2C12 myoblasts were cotransfected with mCherry, mCherry-SQSTM1-WT, or mCherry-SQSTM1 with 1 of 3 different disease mutations (PL, MV, or AX) and GFP-TIA1-WT or 1 of 2 variants (EK or NS) incubated at 42°C for 1 hour and subsequently returned to 37°C for the indicated durations. (E) Bar graph of LDH release from C2C12 myoblasts similar to those in D, before HS and after 1 hour of HS, with an additional 1-hour recovery at 37°C. The absorbance of the samples was measured at 492 nm. The reference wavelength at 680 nm was measured. Representative data were pooled from 3 independent experiments (n = 150~200). Error bars represent the mean ± SEM. *P < 0.05, by 2-way ANOVA and 2-tailed Student’s t test.