A serum microRNA sequence reveals fragile X protein pathology in amyotrophic lateral sclerosis

Abstract

Knowledge about converging disease mechanisms in the heterogeneous syndrome amyotrophic lateral sclerosis (ALS) is rare, but may lead to therapies effective in most ALS cases. Previously, we identified serum microRNAs downregulated in familial ALS, the majority of sporadic ALS patients, but also in presymptomatic mutation carriers. A 5-nucleotide sequence motif (GDCGG; D = G, A or U) was strongly enriched in these ALS-related microRNAs. We hypothesized that deregulation of protein(s) binding predominantly to this consensus motif was responsible for the ALS-linked microRNA fingerprint. Using microRNA pull-down assays combined with mass spectrometry followed by extensive biochemical validation, all members of the fragile X protein family, FMR1, FXR1 and FXR2, were identified to directly and predominantly interact with GDCGG microRNAs through their structurally disordered RGG/RG domains. Preferential association of this protein family with ALS-related microRNAs was confirmed by in vitro binding studies on a transcriptome-wide scale. Immunohistochemistry of lumbar spinal cord revealed aberrant expression level and aggregation of FXR1 and FXR2 in C9orf72- and FUS-linked familial ALS, but also patients with sporadic ALS. Further analysis of ALS autopsies and induced pluripotent stem cell-derived motor neurons with FUS mutations showed co-aggregation of FXR1 with FUS. Hence, our translational approach was able to take advantage of blood microRNAs to reveal CNS pathology, and suggests an involvement of the fragile X-related proteins in familial and sporadic ALS already at a presymptomatic stage. The findings may uncover disease mechanisms relevant to many patients with ALS. They furthermore underscore the systemic, extra-CNS aspect of ALS.

Effects of the investigational drug sodium phenylbutyrate-TUDCA (AMX0035) on the transcriptional and metabolic landscape of sporadic ALS fibroblasts

“…We identified several genes connected to RNA pol II transcription strongly downregulated by Combo, including YBX1 and FXR1 . RNA pol II transcription-associated DNA damage is increased in ALS (83), and these two specific genes are dysregulated and affect RNA homeostasis and transcriptional activity in sALS (84, 85). Lastly, Combo modulates innate immune pathways involving cGAS/STING signaling, which has been reported to be activated in ALS (42).…”

FXR1-related congenital myopathy: expansion of the clinical and genetic spectrum

“…Both FXR1 and FXR2 were found to interact directly with ALS-related microRNAs via GDCGG motif-specific binding, and FXR1 was found specifically to co-aggregate with FUS in motor neurons. This implies a broader role for the family of Fragile X-related proteins in neuromuscular diseases beyond structural myopathies 10. Variants in FXR1 have been associated with several neuropsychiatric diseases, notably insomnia, bipolar disorder and schizophrenia 11 12.…”

RNA Deregulation in Amyotrophic Lateral Sclerosis: The Noncoding Perspective

“…Again, a whole transcriptome profiling, combined with subcellular fractionation analysis of NSC-34 human SOD1 cells, revealed that miR-18b-5p heads a complex gene pathway made up of Hif1α, Mef2c, miR-206, Mctp1, and Rarb, with a downstream effect on cell apoptosis [ 102 ]. A very original contribution recently came from Freischmidt and colleagues [ 103 ] who identified, by proteomic and biochemical studies, the members of the fragile X protein family as interactors of a short sequence motif, enriched in a signature of previously identified ALS-related miRNAs [ 104 , 105 ].…”

“…Again, a whole transcriptome profiling, combined with subcellular fractionation analysis of NSC-34 human SOD1 cells, revealed that miR-18b-5p heads a complex gene pathway made up of Hif1α, Mef2c, miR-206, Mctp1, and Rarb, with a downstream effect on cell apoptosis [102]. A very original contribution recently came from Freischmidt and colleagues [103] who identified, by proteomic and biochemical studies, the members of the fragile X protein [106], an RBP implicated in neural function and degeneration [107], and of proapoptotic targets such as p53. In concordance with this result, in the sALS wobbler mouse-a model displaying almost all clinical hallmarks of human ALS patients [108]-miRNA-375-3p downregulation resulted in inefficient p53 inhibition, increased production of reactive oxygen species, and induced apoptosis [109].…”