How dipeptide repeats cause pathology: A repeat expansion in the chromosome 9 open reading frame 72 ( C9orf72 ) gene is the most common known cause of two neurodegenerative diseases: frontotemporal dementia and amyotrophic lateral sclerosis. This expansion leads to the abnormal production of proteins of repeating dipeptides, but their contribution to disease pathogenesis remains unclear. Zhang et al. engineered a mouse model to study the consequences of one of these dipeptides—prolinearginine dipeptide repeat protein, poly(PR)—in the brain. They found that poly(PR) caused neuron loss as well as motor and memory impairments. These detrimental effects resulted from poly(PR)-induced perturbation of heterochromatin function, a tightly packed form of DNA that represses gene expression. Science , this issue p. eaav2606

Liquid-liquid phase separation of RNA-binding proteins mediates the formation of numerous membraneless organelles with essential cellular function. However, aberrant phase transition of these proteins leads to the formation of insoluble protein aggregates, which are pathological hallmarks of neurodegenerative diseases including ALS and FTD. TDP-43 and FUS are two such RNA-binding proteins that mislocalize and aggregate in patients of ALS and FTD. They have similar domain structures that provide multivalent interactions driving their phase separation in vitro and in the cellular environment. In this article, we review the factors that mediate and regulate phase separation of TDP-43 and FUS. We also review evidences that connect the phase separation property of TDP-43 and FUS to their functional roles in cells. Aberrant phase transition of TDP-43 and FUS leads to protein aggregation and disrupts their regular cell function. Therefore, restoration of functional protein phase of TDP-43 and FUS could be beneficial for neuronal cells. We discuss possible mechanisms for TDP-43 and FUS aberrant phase transition and aggregation while reviewing the methods that are currently being explored as potential therapeutic strategies to mitigate aberrant phase transition and aggregation of TDP-43 and FUS.

Mutations in HNRNPA1 encoding heterogeneous nuclear ribonucleoprotein (hnRNP) A1 are a rare cause of amyotrophic lateral sclerosis (ALS) and multisystem proteinopathy (MSP). hnRNPA1 is part of the group of RNA-binding proteins (RBPs) that assemble with RNA to form RNPs. hnRNPs are concentrated in the nucleus and function in pre-mRNA splicing, mRNA stability, and the regulation of transcription and translation. During stress, hnRNPs, mRNA, and other RBPs condense in the cytoplasm to form stress granules (SGs). SGs are implicated in the pathogenesis of (neuro-)degenerative diseases, including ALS and inclusion body myopathy (IBM). Mutations in RBPs that affect SG biology, including FUS, TDP-43, hnRNPA1, hnRNPA2B1, and TIA1, underlie ALS, IBM, and other neurodegenerative diseases. Here, we characterize 4 potentially novel HNRNPA1 mutations (yielding 3 protein variants: *321Eext*6, *321Qext*6, and G304Nfs*3) and 2 known HNRNPA1 mutations (P288A and D262V), previously connected to ALS and MSP, in a broad spectrum of patients with hereditary motor neuropathy, ALS, and myopathy. We establish that the mutations can have different effects on hnRNPA1 fibrillization, liquid-liquid phase separation, and SG dynamics. P288A accelerated fibrillization and decelerated SG disassembly, whereas *321Eext*6 had no effect on fibrillization but decelerated SG disassembly. By contrast, G304Nfs*3 decelerated fibrillization and impaired liquid phase separation. Our findings suggest different underlying pathomechanisms for HNRNPA1 mutations with a possible link to clinical phenotypes.

Arginine-rich dipeptide repeat proteins (R-DPRs), abnormal translational products of a GGGGCC hexanucleotide repeat expansion in C9ORF72 , play a critical role in C9ORF72 -related amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), the most common genetic form of the disorders (c9ALS/FTD). R-DPRs form liquid condensates in vitro, induce stress granule formation in cultured cells, aggregate, and sometimes coaggregate with TDP-43 in postmortem tissue from patients with c9ALS/FTD. However, how these processes are regulated is unclear. Here, we show that loss of poly(ADP-ribose) (PAR) suppresses neurodegeneration in c9ALS/FTD fly models and neurons differentiated from patient-derived induced pluripotent stem cells. Mechanistically, PAR induces R-DPR condensation and promotes R-DPR–induced stress granule formation and TDP-43 aggregation. Moreover, PAR associates with insoluble R-DPR and TDP-43 in postmortem tissue from patients. These findings identified PAR as a promoter of R-DPR toxicity and thus a potential target for treating c9ALS/FTD.