(A–F and I) Bulk RNA sequencing was performed on cell-sorted pDCs from HNSCC patients (n = 7) (Supplemental Table 2). DEGs were determined using the criterion of fold changes (FC) greater than or equal to 1.5 (see Methods). (A) Principal component analysis showing the clustering of transcriptional profiles of TME and non-TME pDCs (top), including pDCs from HPV⁺ patients (bottom). (B) Top: log₂ expression of TNF-receptor genes upregulated in the TME of pDC samples. Bottom: expression of TNFSF4 in the TME versus non-TME pDCs. (C) Volcano plot of gene expression in OX40⁺ pDCs relative to their expression in OX40^lo/– pDCs (up- and downregulated in OX40⁺ pDCs colored as magenta and orange, respectively) against the FDR. (D) Top pathways (–log[P value] > 2.0) generated from IPA enriched in OX40⁺ and OX40^lo/– pDCs, respectively. Corresponding ratios for each enriched pathway are measured on the second y axis. Full pathway title: Nitrous Oxide and Reactive Oxygen Species Production in Macrophages. (E) Expression of genes involved in MHC I antigen presentation enriched in HPV⁺ HNSCC OX40⁺ pDCs. (F) Heat map reporting relative expression of 183 DEGs in OX40⁺/OX40^lo/– pDCs. Gene pathway clusters are demarcated by boxes. (G) Functional correlative data for cluster II (OXPHOS) in OX40⁺ pDCs. Left: Mitochondrial mass measurements in pDC subsets from the TME were calculated using Imaris (see Methods). Original magnification, ×100. Scale bar: 5 μm. n = 3; 3 experimental repeats. Right: Real-time analysis of OXPHOS (oxygen consumption ratios measured by Seahorse assay) in sorted pDCs from the TME. n = 2; 2 experimental repeats. (H) Functional correlative data for cluster III (detoxification of ROS): total ROS volumes and ROS colocalized to mitochondria volumes in pDCs from the TME. n = 2; 2 patient experimental repeats. (I) Expression of genes involved in cluster VI (type I IFN signaling) upregulated in OX40⁺ pDCs. Unpaired Student’s t test (B and F–I) and right-tailed Fisher’s exact test (E). *P < 0.05; **P < 0.01, ***P < 0.001; NS, not significant. Error bars represent mean ± SEM of technical duplicates. Middle line of box-and-whisker plot indicates the median, box limits indicate the first and third quartiles, and whiskers indicate “extreme” for all data points. Z scores and genes for IPA in Source Data 1.