Journal: bioRxiv
Article Title: Proteomic and in silico dissection of MetaAggregates in amyotrophic lateral sclerosis brains
doi: 10.1101/2025.07.09.663594
Figure Lengend Snippet: (A) A combined proteomic and transcriptomic strategy was employed to search for tripartite MetaAggregates composed of RNA, RBP, and amyloids in ALS brains. Single-cell RNA-seq data from iPSC-derived motor neurons were analyzed to identify disease-associated ELAVL4 and biotinylated rG4-containing RNA sequences (IGFBP2-rG4). A synthetic photo-crosslinkable probe mimicking the tripartite MetaAggregate was generated using psoralen and diazirine chemistry, and used to pull-down binding proteins via NeutrAvidin beads. (B) In the workflow, the probe was incubated with brain lysates from three ALS patients or three healthy controls to capture the associated aggregative proteins. Protein identities were subsequently determined via LC-MS/MS. PCA incorporating amyloidogenicity-related scores was conducted to prioritize candidate proteins, followed by docking simulations with ELAVL4 to assess their structural stability and interaction interfaces.
Article Snippet: A biotinylated 35-nt RNA sequence from IGFBP2 in D-PBS(−) (GGGGGAGGAAGGGGGUU-GUGGUCGGGGAGCUGGGG; IGFBP2-rG4, FASMAC, Kanagawa, Japan) was added to the fraction containing His-ELAVL4.
Techniques: RNA Sequencing, Derivative Assay, Generated, Binding Assay, Incubation, Liquid Chromatography with Mass Spectroscopy