Post-translational modifications (PTMs) affecting proteins during or after their synthesis play a crucial role in their localization and function. The modification of these PTMs under pathophysiological conditions, i.e., their appearance, disappearance, or variation in quantity caused by a pathological environment or a mutation, corresponds to post-translational variants (PTVs). These PTVs can be directly or indirectly involved in the pathophysiology of diseases. Here, we present the PTMs and PTVs of four major amyotrophic lateral sclerosis (ALS) proteins, SOD1, TDP-43, FUS, and TBK1. These modifications involve acetylation, phosphorylation, methylation, ubiquitination, SUMOylation, and enzymatic cleavage. We list the PTM positions known to be mutated in ALS patients and discuss the roles of PTVs in the pathophysiological processes of ALS. In-depth knowledge of the PTMs and PTVs of ALS proteins is needed to better understand their role in the disease. We believe it is also crucial for developing new therapies that may be more effective in ALS.

Pathogenic variations in the fused in sarcoma (FUS) gene are associated with rare and aggressive forms of amyotrophic lateral sclerosis (ALS). As FUS-ALS is a dominant disease, a targeted, allele-selective approach to FUS knockdown is most suitable. Antisense oligonucleotides (AOs) are a promising therapeutic platform for treating such diseases. In this study, we have explored the potential for allele-selective knockdown of FUS. Gapmer-type AOs targeted to two common neutral polymorphisms in FUS were designed and evaluated in human fibroblasts. AOs had either methoxyethyl (MOE) or thiomorpholino (TMO) modifications. We found that the TMO modification improved allele selectivity and efficacy for the lead sequences when compared to the MOE counterparts. After TMO-modified gapmer knockdown of the target allele, up to 93% of FUS transcripts detected were from the non-target allele. Compared to MOE-modified AOs, the TMO-modified AOs also demonstrated reduced formation of structured nuclear inclusions and SFPQ aggregation that can be triggered by phosphorothioate-containing AOs. How overall length and gap length of the TMO-modified AOs affected allele selectivity, efficiency and off-target gene knockdown was also evaluated. We have shown that allele-selective knockdown of FUS may be a viable therapeutic strategy for treating FUS-ALS and demonstrated the benefits of the TMO modification for allele-selective applications.

Wasteosomes (or corpora amylacea) are polyglucosan bodies that appear in the human brain with aging and in some neurodegenerative diseases, and have been suggested to have a potential role in a nervous system cleaning mechanism. Despite previous studies in several neurodegenerative disorders, their status in frontotemporal lobar degeneration (FTLD) remains unexplored. Our study aims to characterize wasteosomes in the three primary FTLD proteinopathies, assessing frequency, distribution, protein detection, and association with aging or disease duration. Wasteosome scores were obtained in various brain regions from 124 post-mortem diagnosed sporadic FTLD patients, including 75 participants with tau (FTLD-tau), 42 with TAR DNA-binding protein 43 (FTLD-TDP), and 7 with Fused in Sarcoma (FTLD-FUS) proteinopathies, along with 29 control subjects. The wasteosome amount in each brain region for the different FLTD patients was assessed with a permutation test with age at death and sex as covariables, and multiple regressions explored associations with age at death and disease duration. Double immunofluorescence studies examined altered proteins linked to FTLD in wasteosomes. FTLD patients showed a higher accumulation of wasteosomes than control subjects, especially those with FTLD-FUS. Unlike FTLD-TDP and control subjects, wasteosome accumulation did not increase with age in FTLD-tau and FTLD-FUS. Cases with shorter disease duration in FTLD-tau and FTLD-FUS seemed to exhibit higher wasteosome quantities, whereas FTLD-TDP appeared to show an increase with disease progression. Immunofluorescence studies revealed the presence of tau and phosphorylated-TDP-43 in the periphery of isolated wasteosomes in some patients with FTLD-tau and FTLD-TDP, respectively. Central inclusions of FUS were observed in a higher number of wasteosomes in FTLD-FUS patients. These findings suggest a role of wasteosomes in FTLD, especially in the more aggressive forms of FLTD-FUS. Detecting these proteins, particularly FUS, in wasteosomes from cerebrospinal fluid could be a potential biomarker for FTLD.

BACKGROUND: Helicase for meiosis 1 (HFM1), a putative DNA helicase expressed in germ-line cells, has been reported to be closely associated with premature ovarian insufficiency (POI). However, the underlying molecular mechanism has not been clearly elucidated. The aim of this study was to investigate the function of HFM1 in the first meiotic prophase of mouse oocytes.
RESULTS: The results suggested that the deficiency of HFM1 resulting in increased apoptosis and depletion of oocytes in mice, while the oocytes were arrested in the pachytene stage of the first meiotic prophase. In addition, impaired DNA double-strand break repair and disrupted synapsis were observed in the absence of HFM1. Further investigation revealed that knockout of HFM1 promoted ubiquitination and degradation of FUS protein mediated by FBXW11. Additionally, the depletion of HFM1 altered the intranuclear localization of FUS and regulated meiotic- and oocyte development-related genes in oocytes by modulating the expression of BRCA1.
CONCLUSIONS: These findings elaborated that the critical role of HFM1 in orchestrating the regulation of DNA double-strand break repair and synapsis to ensure meiosis procession and primordial follicle formation. This study provided insights into the pathogenesis of POI and highlighted the importance of HFM1 in maintaining proper meiotic function in mouse oocytes.

Aberrant condensation and localization of the RNA-binding protein (RBP) fused in sarcoma (FUS) occur in variants of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Changes in RBP function are commonly associated with changes in axonal cytoskeletal organization and branching in neurodevelopmental disorders. Here, we asked whether branching defects also occur in vivo in a model of FUS-associated disease. We use two reported Xenopus models of ALS/FTD (of either sex), the ALS-associated mutant FUS(P525L) and a mimic of hypomethylated FUS, FUS(16R). Both mutants strongly reduced axonal complexity in vivo. We also observed an axon looping defect for FUS(P525L) in the target area, which presumably arises due to errors in stop cue signaling. To assess whether the loss of axon complexity also had a cue-independent component, we assessed axonal cytoskeletal integrity in vitro. Using a novel combination of fluorescence and atomic force microscopy, we found that mutant FUS reduced actin density in the growth cone, altering its mechanical properties. Therefore, FUS mutants may induce defects during early axonal development.

Ferroptosis is a new type of programmed cell death, which has been involved in the progression of tumours. However, the regulatory network of ferroptosis in pancreatic cancer is still largely unknown. Here, using datasets from GEO and TCGA, we screened HSPB1, related to the P450 monooxygenase signalling, a fuel of ferroptosis, to be a candidate gene for regulating pancreatic cancer cell ferroptosis. We found that HSPB1 was enriched in the exosomes derived from human pancreatic cancer cell lines SW1990 and Panc-1. Then, hypoxic SW1990 cells were incubated with exosomes alone or together with HSPB1 siRNA (si-HSPB1), and we observed that exosomes promoted cell proliferation and invasion and suppressed ferroptosis, which was reversed by si-HSPB1. Moreover, we found a potential binding affinity between HSPB1 and FUS, verified their protein interaction by using dual-colour fluorescence colocalization and co-IP assays, and demonstrated the promoting effect of FUS on oxidative stress and ferroptosis in hypoxic SW1990 cells. Subsequently, FUS was demonstrated to bind with and stabilize the mRNA of Nrf2, a famous anti-ferroptosis gene that negatively regulates the level of P450. Furthermore, overexpressing FUS and activating the Nrf2/HO-1 pathway (using NK-252) both reversed the inhibitory effect of si-HSPB1 on exosome functions. Finally, our in vivo studies showed that exosome administration promote tumour growth in nude mice of xenotransplantation, which was able to be eliminated by knockdown of HSPB1. In conclusion, exosomal HSPB1 interacts with the RNA binding protein FUS and decreases FUS-mediated stability of Nrf2 mRNA, thus suppressing hypoxia-induced ferroptosis in pancreatic cancer.

Non-invasive brain stimulation (NIBS) is a complex and multifaceted approach to modulating brain activity and holds the potential for broad accessibility. This work discusses the mechanisms of the four distinct approaches to modulating brain activity non-invasively: electrical currents, magnetic fields, light, and ultrasound. We examine the dual stochastic and deterministic nature of brain activity and its implications for NIBS, highlighting the challenges posed by inter-individual variability, nebulous dose-response relationships, potential biases and neuroanatomical heterogeneity. Looking forward, we propose five areas of opportunity for future research: closed-loop stimulation, consistent stimulation of the intended target region, reducing bias, multimodal approaches, and strategies to address low sample sizes.

Increased signs of DNA damage have been associated to aging and neurodegenerative diseases. DNA damage repair mechanisms are tightly regulated and involve different pathways depending on cell types and proliferative vs. postmitotic states. Amongst them, fused in sarcoma (FUS) was reported to be involved in different pathways of single- and double-strand break repair, including an early recruitment to DNA damage. FUS is a ubiquitously expressed protein, but if mutated, leads to a more or less selective motor neurodegeneration, causing amyotrophic lateral sclerosis (ALS). Of note, ALS-causing mutation leads to impaired DNA damage repair. We thus asked whether FUS recruitment dynamics differ across different cell types putatively contributing to such cell-type-specific vulnerability. For this, we generated engineered human induced pluripotent stem cells carrying wild-type FUS-eGFP and analyzed different derivatives from these, combining a laser micro-irradiation technique and a workflow to analyze the real-time process of FUS at DNA damage sites. All cells showed FUS recruitment to DNA damage sites except for hiPSC, with only 70% of cells recruiting FUS. In-depth analysis of the kinetics of FUS recruitment at DNA damage sites revealed differences among cellular types in response to laser-irradiation-induced DNA damage. Our work suggests a cell-type-dependent recruitment behavior of FUS during the DNA damage response and repair procedure. The presented workflow might be a valuable tool for studying the proteins recruited at the DNA damage site in a real-time course.

Amyotrophic lateral sclerosis (ALS) is a rapidly progressive disease that affects motor neurons, leading to paralysis and death usually 3-5 years after the onset of symptoms. The investigation of both sporadic and familial ALS highlighted four main genes that contribute to the pathogenesis of the disease: SOD1, FUS, TARDBP and C9orf72. This study aims to provide a comprehensive investigation of genetic variants found in SOD1, FUS and TARDBP genes in Greek sporadic ALS (sALS) cases. Our sequencing analysis of the coding regions of the abovementioned genes that include the majority of the variants that lead to ALS in 32 sALS patients and 3 healthy relatives revealed 6 variants in SOD1, 19 variants in FUS and 37 variants in TARDBP, of which the SOD1 p.D90A and the FUS c.*356G>A (rs886051940) variants have been previously associated with ALS, while two novel nonsense pathogenic variants were also identified, namely FUS p.R241* and TDP-43 p.Y214*. Our study contributes to the worldwide effort toward clarifying the genetic basis of sALS to better understand the disease\'s molecular pathology.

Amyotrophic lateral sclerosis (ALS) is a common adult-onset neurodegenerative disease characterized by the progressive death of motor neurons in the cerebral cortex, brain stem, and spinal cord. The exact mechanisms underlying the pathogenesis of ALS remain unclear. The current consensus regarding the pathogenesis of ALS suggests that the interaction between genetic susceptibility and harmful environmental factors is a promising cause of ALS onset. The investigation of putative harmful environmental factors has been the subject of several ongoing studies, but the use of transgenic animal models to study ALS has provided valuable information on the onset of ALS. Here, we review the current common invertebrate genetic models used to study the pathology, pathophysiology, and pathogenesis of ALS. The considerations of the usage, advantages, disadvantages, costs, and availability of each invertebrate model will also be discussed.