Fig 2: Tumor assembly of the self-propagating myeloid niche.a Diagram of transplantation strategies of male primary tumor into female NOD SCID mice. b RNAScope images of transplants showing male-specific Uty+ (red) F4/80+ (green) macrophages within the tumor. Yellow arrows point to F4/80+Uty+ cells. Image is representative of n = 2 independent tumors. c Diagram of transplantation strategies and the ability to trace SCAMs from primary tumor using the CD45.2 allelic variant. d Summary of the proportions for the CD45.2- and Cd45.2+ from either Cd11b+ (left) or Cd11b+Trem2+ cells in the P0, P1, and P2 tumors. e Diagram summarizing self-propagating logic with approximate cell numbers. f Flow cytometry for Trem2 cells in the Cd11b+Cd45.2+ fractions from P0 and P1 tumors. g Quantification of the Trem2% of the Cd11b+Cd45.2+ and Cd11b+Cd45.2- form the P0, P1, and P2 tumors. h Diagram of the transplantation strategy and subsequent sorting to isolate Cd45.2+ and CD45.2- cells from the P1 tumors for scRNA-Seq. i UMAP plots showing the different clusters from the Cd45.2+ and CD45.2- cells. j Feature plots showing key genes associated with SCAMs. k Violin plots showing key genes associated with SCAMs. l Diagram of the intratumor monocyte maturation injection strategy and subsequent analysis by flow cytometry. m Flow cytometry analysis of the injected GFP+ monocytes for SCAM-specific markers. n Quantification of the SCAM-specific markers Trem2 and Vcam1 (n = 4 independent injections into 4 different tumors). Length of each scale bar is noted in figure. For d, g, n = 4 tumors from P0, P1, and P2 allografts. Error bars represent mean +/- SD. p values were calculated using unpaired, two-tailed t test. Source data are provided as a Source data file.

Fig 3: Trem2+Vcam1+ skin cancer associated myeloid cells (SCAMs) are distinct from other non-cancer associated Trem2+ myeloid cells.a UMAP of merged scRNA-Seq from our myeloid cells from mBCC (n = 2 primary tumors), normal skin (n = 4 independent samples), and during wound healing (n = 3 independent samples). Clusters: normal (green), wounded macrophages (blue), wounded dendritic cells (purple), and BCCs (red). b Feature plots for Itgax, Itgam, Adgre1, and Trem2. c Marker genes and GO analysis associated with a. d Stacked violin plots showing marker genes associated with clusters in a. e Feature plot of Vcam1 expression for the scRNA-Seq data in a. f Flow cytometry analysis and quantification of VCAM1/Vcam1 in human and mouse BCCs (n = 3 independent tumors). Error bars represent mean +/- SD. g RNAScope confirmation of Trem2 (green) and Vcam1 (magenta) in mBCCs. Magenta arrows indicate dual Trem2+Vcam1+ cells. Scale bar = 50 µm. Image is representative of n = 2 independent tumors. h Feature plot of VCAM1 expression for the scRNA-Seq data in Fig. 3b. i Updated macrophage polarization summary diagram. j Feature plot of Mki67 expression in the scRNA-Seq data in a. k Macrophage proliferation scoring of clusters in a. l Diagram of the sorting strategies for Trem2+ cells from normal skin, wounded skin, and BCCs. m PCA plot of the differential bulk ATAC-Seq analysis for Trem2+ cells form the different tissues (n = 2 samples from each tissue) in l. n DESeq comparisons between the Trem2+ cells form the different tissues in l. o GO terms are associated with the list of genes associated with the differential peaks from DESEq comparisons. p values were calculated from the hypergeometric distribution. p Example tracks for Trem2 and Vcam1 showing open peaks (highlighted in red) that are specific to Trem2+ cells from the mBCC. q Motifs enriched and de-enriched between SCAMs and Trem2+ cells from the normal tissue. Motifs and subsequent p values were derived from the HOMER pipeline, which uses cumulative binomial distribution. r Venn diagram of the AP-1 motif enrichment/de-enrichment at unique and shared sites that correspond to specific genes. Source data are provided as a Source data file.