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John, S. J.

Publications and source records attributed to John, S. J..

2 recordsLinked to original sources

STMN2 protein depletion via translation deficits and stress granules and its compensation in ALS

STMN2 is an abundant neurospecific protein dysregulated in neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS). We previously reported that cellular stress can lead to STMN2 loss due to TDP-43 nuclear condensation. Here, using human and murine neuronal cell models, multiple pharmacological tools, single-molecule in situ analysis of mRNA localisation and translation, and longitudinal analysis of neuronal fitness/survival, we establish TDP-43-independent mechanisms of STMN2 depletion linked to stress response. We find that human STMN2 protein is extremely labile under acute high-magnitude stress. Early in stress, STMN2 is suppressed via activated proteasomal degradation, phosphorylation and translational repression by stress granules, independently of TDP-43 loss of function in splicing. We further show that STMN2 protein level is highly sensitive to chronic translation deficits, such as those elicited by prolonged low-grade stress. Finally, we demonstrate that STMN2 mRNA is upregulated in non-TDP ALS such as ALS-FUS, which may compensate for translation/stress granule defects in these disease subtypes. Consistent with the compensation hypothesis, STMN2 mRNA is also upregulated in the relatively spared (cortex), but not severely affected (spinal cord), CNS regions in ALS-TDP. In conclusion, our study implicates two common hallmarks of neurodegeneration, translation impairment and abnormal stress granules, in STMN2 depletion and reports an RNA-level compensation that fails in neurons with TDP-43 pathology. Our study supports the development of stress response targeting therapies in ALS with and without TDP-43 pathology.

cell biology↗

FUS post-transcriptional splicing is autoregulated via RNA condensation with therapeutic potential for ALS-FUS

Mutations in the FUS gene cause aggressive and often juvenile forms of amyotrophic lateral sclerosis (ALS-FUS). In addition to mRNA, the FUS gene gives rise to a partially processed RNA with retained introns 6 and 7. We demonstrate that these FUSint6&7-RNAs form nuclear condensates scaffolded by the highly structured intron 7 and associated with nuclear speckles. Using hybridization-proximity labelling proteomics, we show that the FUSint6&7-RNA condensates are enriched in splicing factors and the m6A reader YTHDC1. These ribonucleoprotein structures facilitate post-transcriptional FUS splicing and depend on m6A/YTHDC1 for their maintenance. FUSint6&7-RNAs become hypermethylated in cells expressing mutant FUS, leading to their enhanced condensation and consequently, splicing. We further demonstrate that FUS protein is repelled by m6A. Thus, ALS-FUS mutations may cause an abnormal activation of FUS post-transcriptional splicing via altered RNA methylation. Strikingly, ectopic expression of FUS intron 6&7 sequences dissolves the endogenous FUSint6&7-RNA condensates, downregulating FUS mRNA and protein. Overall, we describe an RNA condensation-dependent mechanism regulating FUS splicing that can be harnessed for developing new therapies.

cell biology↗