Fig. 3

Visualization of significantly enriched gene sets different comparisons. (A) Spatial transcriptomics profiling was conducted on an R/M HNSCC tissue sample. Demarcation of the tissue was achieved through masking PanCk + and PanCk- regions to delineate tumor and stromal compartments, respectively. The morphology markers included PanCk (green), CD45 (red), and SYTO 13 (blue) for the tumor cells, immune cells, and nucleus, respectively. Utilizing the CD45+/- feature, masks were generated to liberate barcodes for sequencing via the Illumina’s NovaSeq 6000 platform. (B) The differential gene expression in TLS regions against GCs visualised as mean transcript expression (AveExpr, in log2) versus fold change (logFC, in log2). Color represents markers that are not differentially expressed (gray), significantly upregulated (red), or downregulated (blue), based on a false discovery rate (FDR) of < 0.05 following multiple testing adjustments. (C, E, F, and G) Gene-set clusters identifying dominant biological themes (E-G) upregulated or (C) downregulated in the comparison TLS vs. GCs with gene-set names depicted as representative wordclouds. Clusters 1,2,4, and 6 representing distinct biological themes were identified in each comparative analysis. (D, H, I, and J) The corresponding gene statistics (i.e. fold change in log2) within the gene sets clusters are plotted against the number of gene sets in the cluster to which the gene is differentially expressed