m6A Modified Short RNA Fragments Inhibit Cytoplasmic TLS/FUS Aggregation Induced by Hyperosmotic Stress

Abstract

Translocated in LipoSarcoma/Fused in Sarcoma (TLS/FUS) is a nuclear RNA binding protein whose mutations cause amyotrophic lateral sclerosis. TLS/FUS undergoes LLPS and forms membraneless particles with other proteins and nucleic acids. Interaction with RNA alters conformation of TLS/FUS, which affects binding with proteins, but the effect of m6A RNA modification on the TLS/FUS–RNA interaction remains elusive. Here, we investigated the binding specificity of TLS/FUS to m6A RNA fragments by RNA pull down assay, and elucidated that both wild type and ALS-related TLS/FUS mutants strongly bound to m6A modified RNAs. TLS/FUS formed cytoplasmic foci by treating hyperosmotic stress, but the cells transfected with m6A-modified RNAs had a smaller number of foci. Moreover, m6A-modified RNA transfection resulted in the cells obtaining higher resistance to the stress. In summary, we propose TLS/FUS as a novel candidate of m6A recognition protein, and m6A-modified RNA fragments diffuse cytoplasmic TLS/FUS foci and thereby enhance cell viability.

m6A RNA methylation in brain injury and neurodegenerative disease

“…This contains the nuclear localization signal (NLS), which alters localization and liquid-liquid phase separation (LLPS) of TLS/FUS in affected neurons, resulting in a further shift in the TLS/FUS into cellular aggregates involved in ALS pathogenesis ( 85 ). TLS/FUS binds intensely to m6A-modified RNA fragments, suggesting that TLS/FUS may serve as m6A readers ( 86 ). Additionally, m6A-modified RNA fragments inhibit aggregation of TLS/FUS in cytoplasmic and affect localization of TLS/FUS-interacting proteins, and thus enhancing cell viability ( 86 ).…”

“…TLS/FUS binds intensely to m6A-modified RNA fragments, suggesting that TLS/FUS may serve as m6A readers ( 86 ). Additionally, m6A-modified RNA fragments inhibit aggregation of TLS/FUS in cytoplasmic and affect localization of TLS/FUS-interacting proteins, and thus enhancing cell viability ( 86 ). Better understanding the regulation of m6A-modified RNA fragments in TLS/FUS aggregation may contribute to development of novel ALS therapeutics.…”

Pseudogenes and Liquid Phase Separation in Epigenetic Expression

“…For instance, YTHDF2 can recruit the m 6 A -containing transcript into P-bodies for being degraded (46). Moreover, Translocated in LipoSarcoma/Fused, a nuclear RNA-binding that forms the membrane-less aggregates, is affected by various m 6 A modifications in mediating the liquid phase separate condensates (47). Lee et al show that the m 6 A modification regulates gene expression through liquid phase separate intervention.…”

Effects of Mass Change on Liquid–Liquid Phase Separation of the RNA-Binding Protein Fused in Sarcoma

“…The functions of biomolecular condensates referred to as membraneless organelles (MLOs) include cell signaling, nuclear transcription, RNA splicing and processing, and DNA sensing and damage repair [ 3 , 5 , 6 , 7 , 9 , 10 , 11 , 12 , 13 ]. Importantly, dysregulation of LLPS has been associated with the pathogenesis of neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), and Alzheimer’s, Parkinson’s, and Huntington’s diseases [ 6 , 14 , 15 , 16 , 17 , 18 ]. Although there is no study which can decipher conclusively the cellular and pathologic basis of the diseases, the unifying observation of abnormal protein inclusions in postmortem tissue may suggest that one well-characterized cellular feature of neurodegenerative disease is the deposition of protein aggregates in affected brain regions [ 19 ].…”