Tau polarizes an aging transcriptional signature to excitatory neurons and glia

Cluster refers to the numeric ID assigned by Seurat when FindClusters resolution is set to 2, and the Annotation column notes the cell identity assignment. This table can be used to obtain cell identities of Seurat cluster IDs in the result tables below.

Cluster markers are obtained by MAST differential expression analysis where each cell cluster is compared against all remaining cells. Only genes with a positive log2 fold change are displayed. Log2 fold change = expression fold change between a given cluster and all remaining cells. Pct.1 = percent of cells in the given cluster with non-zero expression of the gene. Pct.2 = percent of the remaining cells (not in the cluster) that have non-zero expression of the gene. Cluster ID = Seurat assigned cluster ID. BH-adjusted p-value = Benjamini–Hochberg-corrected p-value from the MAST differential expression analysis for each cluster.

Cell library metrics from the 10x Genomics Cell Ranger output. Sample = cell library labels. Libraries from the replication experiment are labeled with ‘rep’ behind the final underscore. See also Figure 1—figure supplement 1. Additional details on the data provided in each column can be found in 10x Genomics support materials: https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/output/gex-metrics.

Table of established fly gene expression markers used as references for cell annotation. Genes = marker genes that can be used to identify a cell type. Cell type = Drosophila brain cell subpopulation. Reference = source publication used to obtain the genes.

Analysis of cell abundance changes between elav>tau^R406W and control animals as quantified by DESeq2. In the discovery dataset, the 1, 10, and 20-day timepoints are pooled, such that n = 3 values for each comparison. The replication dataset is comprised of n = 3 elav>tau^R406W and control (elav-GAL4) animals all prepared at day 10. baseMean = mean of normalized cell counts for the given cell cluster across all samples. Log2FoldChange = log2 fold change of elav>tau^R406W vs. control cell counts. lfcSE = standard error of log2 fold change value. Pvalue = p-value from Wald test of the genotype log2 fold change value. Differences in cell count are quantified by negative binomial GLM, such that count ~genotype + age. Padj = adjusted p values using the Benjamini–Hochberg procedure. Experiment = denotes if data is from the discovery or replication analysis.

Tau-induced differentially expressed genes were adjusted for aging by including a covariate in the regression model, based on comparisons of scRNAseq data elav>tau^R406W vs. control (elav-GAL4) at 1, 10, and 20 days. Aging-induced differentially expressed genes are based on comparisons in control (elav-GAL4) flies, including between day 1 (d1) and day 10 (d10), and day 10 vs. day 20 (d20); comparisons are noted in age_comparisons column. The cell cluster being compared is denoted in the cluster column. Avg_logFC is the log2 fold change of gene expression between day 10 vs. day 1 or day 20 vs. day 10; in each entry, the former is the numerator, and the latter is the denominator. For tau vs. control comparisons, the numerator is tau, and the denominator is control. Pct.1 and Pct.2 refer to the percent of cells that have non-zero expression for the given gene in the numerator and denominator, respectively. P_val = uncorrected p-values from the MAST linear regression. Padj = Benjamini–Hochberg-adjusted p-values. Analysis = specifies either ‘control aging’ or ‘tau age-adjusted’ for the respective analyses.

Significantly enriched functional terms based on overrepresentation analysis (ORA) of cell-specific differentially expressed gene sets, including from either (i) aging (controls), (ii) tau age-adjusted (elav>tau^R406W vs. control (elav-GAL4)), or the (iii) ‘tau-specific’ gene set, which is the unique subset of genes from ii not seen in i. Genes used for functional enrichment analysis have a false discovery rate (FDR) < 0.05 in all differential expression analyses, and all functional enrichment terms listed have a hypergeometric FDR < 0.05. Analysis = source of gene set used for functional enrichment (i–iii, above). Age = relevant age groups of source comparison. Cluster = cell cluster source of gene set. Term_id = identifier of enrichment term. term = description of enriched term. FDR = FDR-corrected p-values. Database = database origin of term.

Cross-sectional replication analysis comparing differentially expressed genes in an independent dataset from day 10 (elav >tau^R406W vs. control (elav-GAL4)). The cell cluster being compared is denoted in the cluster column. Avg_log2FC is the log2 fold change of gene expression between tau vs. control comparisons, the numerator is tau, and the denominator is control. Pct.1 and Pct.2 refer to the percent of cells that have non-zero expression for the named gene in the numerator and denominator, respectively. P_val = unadjusted, raw p-values from the MAST linear regression. Padj = Benjamini–Hochberg-adjusted p-values.

Cell-cluster overlaps are quantified between the age-adjusted discovery data (Figure 3—source data 1) and the day 10 cross-sectional replication data (Figure 3—source data 3). Cluster = annotated cell identities or Seurat ID of unannotated clusters. ageAdj_discovery_DEG_n = number of tau-induced differentially expressed genes (FDR < 0.05) in the discovery dataset. d10_CS_replicate_DEG_n = number of differentially expressed genes in the replication dataset. Intersect = number of overlapping differentially expressed genes between results of the two comparisons. percent_of_original = percent of differentially expressed genes in the discovery dataset that is also observed in the replication dataset. Phyper = p-value of hypergeometric tests evaluating whether the number of overlapping genes observed is greater than by chance. tot_genes = total number of unique genes detected in each cell type and shared between datasets used for hypergeometric test.

Cross-sectional analysis of tau-induced changes (elav>tau^R406W vs. control (elav-GAL4)) from the discovery dataset at 1, 10, and 20 days; age = the age being compared. The specific cell cluster being compared is denoted in the cluster column. Avg_log2FC is the log2 fold change of gene expression between tau vs. control comparisons, the numerator is tau, and the denominator is control. Pct.1 and Pct.2 refer to the percent of cells that have non-zero expression for the named gene in the numerator and denominator, respectively. P_val = unadjusted, raw p-values from the MAST linear regression. Padj = Benjamini–Hochberg-adjusted p-values.

Differential expression of the immune response coexpression module (magenta, Mangleburg et al., 2020), based on comparisons of elav>tau^R406W and control (elav-GAL4) animals, adjusting for age. Cluster = cell cluster identity. lrt.pvalues = uncorrected p-values from likelihood ratio test. lrt.padj = Benjamini–Hochberg-adjusted p-value. log2FC = log2 fold change of mean immune module expression between tau and controls for each cell cluster.

183 regulons and member genes are denoted with the (+) notation, indicating that genes within these modules are positively co-expressed.

Regulon expression per cell was defined as the mean expression of regulon member genes. For each cell type, regulon expression across all cells was regressed on genotype [elav>tau^R406W vs. control (elav-GAL4)] and age, and a likelihood ratio test was performed against a reduced model with age only. Cluster = cell identity. Regulons = regulon used in statistical testing. lrt.pvalues = unadjusted p-values from likelihood ratio test. lrt.padj = Benjamini–Hochberg-adjusted p-values. log2FC = log2 fold change of mean regulon expression between tau and control.

Comprehensive list of retained, non-zero coefficients from the elastic net regression models considering expanded list of 2993 predictor variables and cell clusters showing significant, tau-induced reductions in cell abundance. Term = a variable in the elastic net multiple regression. Elastic net coefficient = the coefficient for the specified variable in the final regression model.

snRNAseq data from human postmortem brain tissues (Mathys et al., 2019) was analyzed for differentially expressed genes in AD neuropathological cases versus controls without AD pathology. Cell subcluster (subcluster) labels are as defined by the original publication. Pct.1 and Pct.2 refer to the percent of cells that have non-zero expression for the given gene in the numerator and denominator, respectively. Avg_log2FC is the log2 fold change of gene expression between AD vs. control comparisons; the numerator is AD, and the denominator is control. P_val = unadjusted p-values from the MAST linear regression analysis. Padj = Benjamini–Hochberg-adjusted p-values.