Single-cell Epigenetics of Tau Dementia | Part 3: Exploring Disease-enriched Glial States, Disease-Reactive Microglia and Astrocytes, and Genetic Heritability | Epigenome Technologies

By Stuart P. Atkinson

Diversity and heterogeneity of cellular subtypes in human brain of tauopathies. (A-C) UMAP embedding of subclusters of astrocytes (A), with a heatmap of gene score matrix labeled by log2fc > 1 for marker gene scores (B) and enriched functional terms associated with these marker genes (C). ASC, astrocytes. (D-F) Similar analyses for microglia subclusters. MG, microglia

Understanding Tauopathies Through Single-cell Epigenetic and Transcriptomic Analyses

The Epigenome Technologies Blog brings you the final part of a three-part summary of a recent single-cell epigenetics preprint article from researchers led by Jessica E. Rexach (University of California, Los Angeles), who sought to define cell-type-specific cis-regulatory elements (CREs) via chromatin accessibility (snATAC-seq) and gene expression (snRNA-seq) analysis in single nuclei across 6 cell types and 50 subclasses in samples from brain regions with distinct vulnerabilities in Alzheimers disease (AD), Picks disease (PiD), and progressive supranuclear palsy (PSP) patients to understand the regulatory circuitry of non-coding genetic variants underlying risk-associated cell states (Han et al.).

The authors provide a cross-disorder atlas linking gene regulation, chromatin dynamics, and cellular functions across tau-related disorders to highlight disorder-specific glial states of differential resilience. In doing so, they enhance our understanding of disease regulatory circuits by uncovering epigenomic dynamics and mapping genetic variants to their target through CREs, prioritize genes for validation to inform causal mechanisms and therapeutic strategies by identifying molecular targets linked to polygenic disease risk, enhance our understanding of glial contributions to tauopathies at the single-cell level, and underscore the importance of cross-disorder, cell-specific chromatin profiling in brain regions with moderate pathology.

Diversity and heterogeneity of cellular subtypes in human brain of tauopathies. (G) Gene signatures of astrocyte subclusters (top) and microglia subclusters (bottom), defined by marker gene scores compared within each respective cell type. (H) RNAscope ISH for SOX10 combined with GFAP IHC in human insular tissue from a PSP patient. Representative images show co-localization of SOX10 mRNA and GFAP in a subset of astrocytes (Scale bar: 10 µm). (I) Boxplots displaying the relative abundance of insula ast.C1 (top) and mg.C4 (bottom) across conditions. Changes in cell composition among disease and control groups are modeled using linear regression computed by Limma, adjusted for age and post-mortem interval. *P<0.05. (J) Example tracks showing marker peaks for genes PLP1, SOT1l in mg.C4; RIN3 and TREM2 in mg.C13 and mg.C6; CD48 in mg.C9 and mg.C6; and LGALS3 in mg.C7 and mg.C14. (K) Example tracks showing marker peaks for genes CNTN2, KLK6, and PLP1 in ast.C1; HLADMB in ast.C3; and GFAP in multiple astrocyte subclusters.

Exploring Disease-enriched Glial States, Disease-Reactive Microglia and Astrocytes, and Genetic Heritability

Integrated analysis of accessibility changes, gene activity, GWAS heritability partition, epigenomic stability, and cell-cell interactions identifies disease-associated glia subtypes. (A) PSP-associated chromatin accessibility changes in astrocytes. Bar plots (left) show the number of differentially accessible CREs in PSP across astrocyte subclusters, categorized by upand down-regulation. The right panel shows partitioned disease heritability of dynamic peaks in ast.C1 and ast.C10 (right), displaying LDSC standardized effect size (). FDR *< 0.05; ** <0.005; *** < 0.001. (B) Differentially activated TFs in astrocytes and ast.C1.

Do Variable Disease-enriched Glial States Associated with Specific Neurological Disorders

Analysis of altered cell composition within brain regions across disease states revealed increased total microglia in AD and astrocytes in PiD, and disease-specific and subcluster composition changes in a region-specific manner
Prominent alterations occurred in oligodendrocytes and PLP1+ myelin-related subtypes in PiD, which exhibited pronounced disease-specific changes in subtype composition in inflammatory oligodendrocytes in the insula

Diseases shared alterations in myelin-related subclusters in the insula, but involved different disorder-specific glial types
C1- and C4-subtype astrocytes displayed significant increases in PiD and PSP, while C6-subtype microglia increased in PiD and AD

The observations of increased accessibility of genes associated with myelination in microglia and astrocytes suggest a shared feature of primary and secondary tauopathies

Furthermore, these findings reveal that the insula accumulates pathological signals, with myelin-related microglia and astrocytes increasing in PiD and PSP

Do Specific Transcription Factors Drive Disease-Reactive Microglia and Astrocytes in PiD and PSP

The study further investigated the gene regulatory mechanisms driving PLP1 and other myelin-related genes in tauopathies and explored how cell stress and injury pathways interact with Tau pathology
Transcription factor (TF) analysis revealed the significant enrichment of SOX10 (regulator of myelination genes) in C4-subtype microglia and C1-subtype astrocytes, distinguishing these subtypes from other astrocytes and microglia
Identifying TF targets mediating binding events in accessible chromatin predicted binding sites on the marker peaks of subclusters

34.4% of SOX10 targets overlapped across oligodendrocytes, astrocytes, and microglia
However, the unique prominence of several targets among SOX10 targets in the non-oligodendrocyte cell types supports the shared and distinct effects of SOX10 by cell type

SOX10 shared targets with the TF FOXP2 (linked to neuropsychiatric disorders) in C4-subtype microglia but shared targets with NFIX (an astrocyte-specific TF) in C1-subtype astrocytes
SOX10 target genes in C4-subtype microglia and C1-subtype astrocytes displayed enrichment in processes, suggesting that these clusters promoted cell growth and lipid metabolism to maintain brain function

C1-subtype astrocytes highly expressed MAPT(encoding tau) compared to all other astrocytes in PSP

Identifying TF target genes in each disorder in C1-subtype astrocytes revealed the differential activation of SOX10 in the PSP insula in all disease conditions

Data show that SOX10 regulates myelin genes/pathways related to Tau toxicity in C1-subtype astrocytes

SOX10 targets and pathways across and within each cell type support a role in regulating changes in lipid metabolism in the context of disease, providing benefits that would support cell survival under stress

Epigenomic erosion analysis revealed significant gains and losses of heterochromatin across multiple astrocyte states in PSP, where C1-subtype astrocytes remained unaffected, suggesting that the maintenance of epigenomic stability may protect them from depletion as a resilient state

Is Genetic Heritability Enriched in Disorder-Divergent Cell States in PSP and PiD

The authors applied linkage disequilibrium score regression to cell subclusters representing distinct states to assess relationships between gene regulation, enhancer co-accessibility, disease risk loci, and downstream pathways using diverse disease contexts

This analysis revealed two clusters across all cell types enriched for PiD heritability and implied five neuron subclusters and two astrocyte clusters as enriched for PSP heritability
Excitatory neurons demonstrated a loss of chromatin accessibility at PSP genome-wide association study (GWAS) loci, while astrocytes exhibited a gain in accessibility at these loci

The authors identified cell states with aggregated co-regulatory structures affecting potential causal SNPs

Differentially accessible CREs (DA-CREs) concentrated in glial subclusters across all disorders/brain regions
C1-subtype astrocytes displayed the most significant number of disease-specific CREs that gained chromatin accessibility in PSP samples in the insula
A significant enrichment of PSP heritability in C1-subtype astrocyte-specific peaks and disease-dynamic peaks suggested these cells as a state harboring PSP heritability-relevant chromatin accessibility
SOX10 was a marker gene for C1-subtype astrocytes, but also exhibited differential activation in PSP

The study prioritized functional genes for C1-subtype astrocytes based on their linked PSP differentially-activated CREs associated with GWAS variants

The findings suggested that C1-subtype astrocytes may enhance intracellular trafficking, including endocytosis and vesicle-mediated transport, in response to tau and myelin accumulation, which may contribute to maintaining protein homeostasis and preventing cellular dysfunction, potentially mitigating neurodegenerative processes in PSP

Analysis of microglia revealed the highest gain of PID heritability in C4-subtype microglia, suggesting that accessibility changes in this subtype might capture PiD causality

Known PID disease modifier genes affecting TDP43 forms of the disease displayed more differential expression in C4-subtype microglia than any other microglia type
One CRE in C4-subtype microglia contained a PiD SNP targeted gene,CTSC, whose product degrades proteins in a lysosome pH-dependent manner

The combination of CRE with linked PID GWAS-associated single-nucleotide polymorphisms (SNPs) and pathway analysis defined downstream biology

Genes differentially activated in PiD in C4-subtype microglia suggested a potential role in regulating microglial pathways related to stress and immune responses

Overall, C4-subtype microglia might be subject to additional lipid regulation, which enhances ER stress responses and activates degradation pathways
Inferred cell-cell interactions revealed higher numbers involved in C4-subtype microglia in PiD compared to the other conditions, suggesting increased cellular interaction activity in PiD

These cell-cell interactions orchestrated single-cell functions to maintain homeostasis and regulate physiological processes

Analysis of interactions between C4-subtype microglia and representative neuronal and glial subclusters reveals key ligand-receptor pairs mediating signaling related to myelination, inflammation, and synaptic regulation, including PiD-risk genes, which further underscores this subcluster as a key subcluster responsive to PiD pathology

What Can Paired-Tag from Epigenome Technologies Do for Your Research?

Paired-Tag from Epigenome Technologies generates joint epigenetic and transcriptomic profiles at single-cell resolution and detects histone modifications and RNA transcripts in individual nuclei with comparable efficiency to single-nucleus RNA-seq/ChIP-seq assays while avoiding the need for cell sorting. The implementation of Paired-Tag technology may enable researchers to make significant strides in understanding gene regulation and improving the management of diseases, such as the neurodegenerative tauopathies explored in this exciting preprint.