Extended Data Fig. 2: Single nucleus ATAC sequencing.

a. Experimental models: All ATAC-seq experiments examining LPS or SCI treatments or healthy controls used wild-type or transgenic mice of 657Bl6 background strain. Transgenic mice expressing mGFAP-Cre¹² were used for astrocyte-specific deletion (cKO) of Smarca4 or Stat3. b. The same region of thoracic spinal cord at T9-T10 was harvested for all evaluations. Spinal cord tissue from different experiments were frozen until all experiments were completed. All tissue was then processed at the same time to limit technical variations. c. Two spinal cords from the same experimental group were pooled to prepare suspensions of nuclei. These suspensions were enriched for astrocyte nuclei with a Sox9 antibody and magnetic beads precipitation. Two such suspensions were prepared from a total of n = 4 mice per experimental condition. d. Two biological replicates, each consisting of nuclei from two mice, were used for single nucleus ATAC-seq. All biological replicates from all experimental conditions were sequenced at the same time to avoid batch effects. e. Box plots compare the distribution of the per-nucleus averages of unique DNA fragments per nucleus or TSS enrichment per nucleus for the seven experimental conditions listed, with each dot representing the average value of all nuclei collected from each experimental group (whiskers show range, box encompass 25–75% quartiles and the centre line indicates the median; n = 8 WT healthy mice and n = 4 mice for all other experimental conditions). f. UMAP clustering based on differential ATAC peaks across 145,973 high quality nuclei isolated across all conditions showed separation of nuclei into multiple distinct clusters. g. UMAP distribution of examples of specific DAGs used to identify the dominant cell types in different clusters. h. Heatmap of DAGs used to identify specific cell types. Each line represents the per-nucleus z-score for a given gene averaged across all nuclei in a cluster. i. UMAP and stacked bar graph show relative contribution of biological replicates for different experimental conditions. Because experiments for LPS and SCI were conducted at different times, groups of healthy control mice were collected for each experiment. All biological replicates showed similar contributions to their respective clusters, confirming that separation of treatment groups with essentially no overlap of LPS with either healthy or SCI was due to biological variation and not due to technical artifacts. * each biological replicate consisted of nuclei from two mice.

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