Main
Results
Cohort of H3-K27M DMGs across age groups and locations
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H3-K27M DMG cell composition across age and location
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Location specificity of OPC-like subpopulations
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The open chromatin landscape of H3-K27M DMG cell populations
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The age-specific myeloid cell landscape in H3-K27M DMGs
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Charting the single-cell spatial architecture of H3-K27M DMG
a, Schematic of HybISS experimental approach. Briefly, mRNA is amplified in situ by RT, and the product cDNA is hybridized with a custom complementary padlock probe. Next, RCA reaction is run to generate a blob of DNA that can then be barcoded with individualized gene bridge probes and fluorescently barcoded. After imaging, the sample is stripped of bridge probes, and the cycle is repeated five times with different fluorophores for decoding and identification of gene signals based on their decoding sequence. b
We next performed neighborhood enrichment analyses to investigate spatial relationships between individual cell populations. Here we observed marked variability in neighborhood structures, highlighting overall intertumoral spatial heterogeneity (Supplementary Fig. 3
Discussion
First, we resolve pontine H3-K27M DMGs to harbor more immature pre-OPC-like tumor cells than their thalamic counterparts. This raises the question of whether this diversity reflects region-specific cell-intrinsic features or it is driven by local environmental interactions. While normal murine OPCs have been shown to lack heterogeneity across different brain regions44,55