Fig. 1
Experimental setup and workflow for circulomics and proteomics profiling in spinal cord. A Total DNA was purified from cervical myelon of control and symptomatic hSOD1G93A mutant animals. B Circular DNA purged from linear DNA was amplified and assembled. C. EccDNA was subjected to short-read sequencing and mapped to genomic coordinates. D–E Computational quantitative algorithms were generated that in silico reconstructed circle sizes, determined the originating genomic coordinates and profiled them as being common or different in each group. Moreover, algorithms were designed to find full gene and telomere-specific circles. Filter means excluded mitochondrial circular DNA. F–G Mass spectrometry-based proteomic studies were performed on cervical spinal cord in a cell compartment-specific manner, dissecting 5 subcellular fractions. Changes in protein abundances between ALS and control specimens, as exemplified in the heatmap for the cytoplasmic (CP) fraction, were selected for eccDNA-related gene products. H Output genes from eccDNA and proteome analyses were referenced to GWAS. For eccDNA analyses, n = 9 for ALS and n = 10 for control samples. For MS studies, n = 5 for both groups. ALS, hSOD1G93A mutant samples; Ctrl, control samples