Module 2, with lower expression in astrocytes from those with AUD, contained 24 transcription factors and 451 target genes. To determine which of these genes were more likely to be regulated together, we performed weighted gene co-expression network analysis (WGCNA48) of astrocyte single-nucleus data, which revealed ten groups of co-expressed genes (Supplementary Table 15). Co-expression group 3 (Co.E3) was enriched for differentially expressed genes, particularly those with lower expression in individuals with AUD. There were 75 genes in Co.E3 that overlapped with regulatory module 2 (Fig. 6B), suggesting that these genes not only show similar expression patterns but also have similar patterns of regulation. Functional enrichment analysis identified several enriched pathways, including pathways relating to glutamatergic synapses, nervous system development, and negative regulation of Wnt signaling pathways from GO Biological Process and Cellular Component (Fig. 6C).
