Extended Data Fig. 3: Stat6 genetic deletion in TAMs.

a) Macrophages transduced with the CRISPR-Cas9 GeCKOv2 library were sorted based on MHC II and CD206 expression. Negative regulators were ranked based on pvalue (<=-0.005) and the first 200 genes were grouped into functional families. The pie chart shows the percentage distribution of the identified genes. b) Experimental scheme of in vitro Stat6 genetic deletion on primary bone marrow derived macrophages. Two independent sgRNA (g1 and g2) have been employed. c) Representative FACS plot showing the expression of CD206 and MHCII on LGP (CTRL) and Stat6 KO (g1 and g2) macrophages. d-e) FACS analysis of control (LGP) and STAT6 silenced (g1 and g2) macrophages upon exposure to Pten-/-; Trp53-/- conditioned media: (d) % of ARG1+ (LGP n = 4, g1 n = 4, g2 n = 4) and (e) CD39+ cells (LGP n = 3, g1 n = 3, g2 n = 3) gated on F4/80 + CD11b+ cells. f) RT-qPCR gene expression analysis on LGP or Stat6 KO macrophages exposed to Pten-/-; Trp53-/- media. Bar graphs show the fold change of treated cells versus untreated for each condition g) Scratch assay: graph and curves showing the distance (um) covered by tumor cells over time after exposure to supernatant from untreated macrophages (n = 9/group). h) Volcano plot showing chemosensor genes related to the differentially enriched sgRNA guides from CD206-MHCII+ vs CD206brightMHCII- cells. Olfactory receptors are shown in orange, Vomeronasal receptors are shown in green. Statistical analyses were performed using two-tailed unpaired Student’s t test. Values are presented as mean ± SEM. Schematic in b created using BioRender.com.