Hodiset al.,Science 376 , eabi8175 (2022) 29 April 2022 9 of 14
ACGB CAF, contractile CAF, immunom. M1 macrophageM1 and M2 macrophagesHI
Complement / Ribosomal S100 / InterferonM1 – most used program
Metabolism / Lgals M2 (cells with M2 program <0.45)M2 macrophage Neutrophil020406080Percent of all stromal cells in a sample (%)Endothelial Epithelial DC pDC Pericyte0 02468510152025UMAP 1UMAP 2CBTP
CBTP (6 mo.)
CBTA
CBTP3
CBTPACBTP
CBTP (6 mo.)
CBTA
CBTP3
CBTPAUMAP 1UMAP 1UMAP 1UMAP 1UMAP 1UMAP 1UMAP 1UMAP 1
FractionUMAP 1CBTA rep. 1
CBTA rep. 2
CBTA rep. 3
CBTP rep. 1
CBTP rep. 2
CBTP rep. 3 #
CBTP (6 mo.) rep. 1
CBTP (6 mo.) rep. 2
CBTP3 rep. 2CBTP3 rep. 1
CBTP3 rep. 3
CBTPA rep. 1
CBTPA rep. 2
CBTPA rep. 3
CAF,
contractileEndothelial NeutrophilsDCM2 macrophagesM1 macrophagespDC Epithelial
PericytesUMAP 2UMAP 2Mutant melanocytes
cultured in vitroInjection of cells of a single genotype
into dermis of immunodeficient mouse36–151 daysTumor growth Single cellsisolation Dissociationof cells scRNA-Seq stromal cellsAnalysis:CBTP rep. 3 #
CBTP rep. 2
CBTP rep. 1
CBTA rep. 3
CBTA rep. 2
CBTA rep. 1
CBTP3 rep. 3CBTP3 rep. 2
CBTP3 rep. 1
CBTPA rep. 3
CBTPA rep. 2
CBTPA rep. 1M1: Compl./Ribo.CBTP rep. 2
CBTP rep. 1(6 mo.)CBTPA rep. 1
CBTPA rep. 2
CBTPA rep. 3CBTP3 rep. 2
CBTP3 rep. 1
CBTP3 rep. 3CBTP (6 mo.) rep. 1CBTP (6 mo.) rep. 2CBTP rep. 1
CBTP rep. 2
CBTP rep. 3 #CBTA rep. 1
CBTA rep. 2
CBTA rep. 3
0.00 0.25 0.50 0.75 1.00M1: S100/Interf.
M2M1: Metab./LgalsDENeutrophils (N) Neutrophils (N) F Neutrophils (N)UMAP 1 UMAP 1 UMAP 1N Sig. 5UMAP 1UMAP 2 UMAP 2
Signature scoreCBTA rep. 3
CBTA rep. 2
CBTA rep. 1
CBTP3 rep. 3
CBTP3 rep. 2
CBTP3 rep. 1–0.250.000.250.500.750.8
0.6
0.4
0.2
0.81.001.25CBTA rep. 1CBTA rep. 2CBTA rep. 3CBTP3 rep. 1CBTP3 rep. 2CBTP3 rep. 3CBTA rep. 1CBTA rep. 2CBTA rep. 3CBTP3 rep. 1CBTP3 rep. 2CBTP3 rep. 3N Sig. 1&3 N Sig. 5 N Sig. 1&30Signature score Signature score
0 0.8 0.4Program 1 Program 2 Program 3 Program 4CAF,
immuno-
modulatory1.0
0.8
0.6
0.4
0.2
0.0Program
usageFig. 5. Tumor genotype shapes tumor microenvironment composition.
(A) Experimental approach to profile gene expression from mouse cells in tumor
xenografts with scRNA-seq. (BandC) Remodeling of the tumor microenvironment
cellular composition by cancer cell genotype and duration in mice. (B) UMAP
embedding of tumor microenvironment scRNA-seq profiles (dots) colored by
genotype and replicate (legend) and labeled by cell type. #, CBTP rep. 3 is a mixture
of four tumors from two mice, whereas all other replicates are from a single tumor.
(C) Mean (bar) and individual (dots) percent (yaxis) of tumor microenvironment
cells of each type (xaxis) in each genotype (color). (DtoF) Diversity of neutrophil
expression programs in tumors of different genotypes. (D) UMAP embedding of
neutrophil single-cell profiles (dots) from specific tumor genotypes (colored) and all
other genotypes (gray). (E) UMAP embedding of neutrophil profiles (as in D)
highlighting only the neutrophils in CBTA and CBPT3 tumors colored by per-cell
score of neutrophil expression signatures, N5 (associated with tumor growth),
and N1N3 (more similar to circulating and healthy-tissue neutrophils); signatures
previously described ( 55 ). (F) Distribution of per-cell neutrophil expression
program score (yaxis) in neutrophils from CBTA (green) and CBTP3 (blue) tumors.
Pvalue < 0.001 (N5),Pvalue < 0.001 (N1N3), Kruskal-Wallis rank-sum test,
degrees of freedom = 5. (GtoI) Effect of tumor genotype on macrophage expression
programs. (G and H) UMAP embedding of M1 and M2 macrophage single-cell profiles
(dots), colored by specific tumor genotypes (G) or by per-cell relative usage of
macrophage gene expression cNMF programs (H). (I) Fraction of cells (xaxis) with
the highest score in each of four cNMF programs (colors) among M1 cells (defined
as cells with an M2-related program score <0.45) in each replicate (yaxis).RESEARCH | RESEARCH ARTICLE