In addition, MSS-A and MSS-N tumors were associated with somatic mutations in different driver genes, with high frequencies of mutated TP53 in MSS-A and mutated KRAS and BRAF in MSS-N. Our results suggest that MSS-A and MSS-N represent two subtypes of MSS colorectal cancer, and such stratification may be used to improve therapeutic treatment in an individualized manner. Defactinib supplier SIGNIFICANCE This study shows that chromosome 20q amplification occurs predominately in microsatellite-stable colorectal cancer and defines a distinct subtype with good prognosis, high chromosomal instability, distinct mutation profiles, and low immune infiltrations.Macrophages are critical mediators of tissue homeostasis, cell proliferation, and tumor metastasis. Tumor-associated macrophages (TAM) are generally associated with tumor-promoting immunosuppressive functions in solid tumors. Here, we examined the transcriptional landscape of adaptor molecules downstream of Toll-like receptors in human cancers and found that higher expression of MYD88 correlated with tumor progression. In murine melanoma, MyD88, but not Trif, was essential for tumor progression, angiogenesis, and maintaining the immunosuppressive phenotype of TAMs. In addition, MyD88 expression in myeloid cells drove melanoma progression. The MyD88/IL1 receptor (IL1R) axis regulated programmed cell death (PD)-1 expression on TAMs by promoting recruitment of NF-κBp65 to the Pdcd1 promoter. Furthermore, a combinatorial immunotherapy approach combining the MyD88 inhibitor with anti-PD-1 blockade elicited strong antitumor effects. Thus, the MyD88/IL1R axis maintains the immunosuppressive function of TAMs and promotes tumor growth by regulating PD-1 expression. SIGNIFICANCE These findings indicate that MyD88 regulates TAM-immunosuppressive activity, suggesting that macrophage-mediated immunotherapy combining MYD88 inhibitors with PD-1 blockade could result in better treatment outcomes in a wide variety of cancers. GRAPHICAL ABSTRACT http//cancerres.aacrjournals.org/content/canres/81/9/2358/F1.large.jpg.Fusion genes including NPM-ALK can promote T-cell transformation, but the signals required to drive a healthy T cell to become malignant remain undefined. In this study, we introduce NPM-ALK into primary human T cells and demonstrate induction of the epithelial-to-mesenchymal transition (EMT) program, attenuation of most T-cell effector programs, reemergence of an immature epigenomic profile, and dynamic regulation of c-Myc, E2F, and PI3K/mTOR signaling pathways early during transformation. A mutant of NPM-ALK failed to bind several signaling complexes including GRB2/SOS, SHC1, SHC4, and UBASH3B and was unable to transform T cells. Finally, T-cell receptor (TCR)-generated signals were required to achieve T-cell transformation, explaining how healthy individuals can harbor T cells with NPM-ALK translocations. These findings describe the fundamental mechanisms of NPM-ALK-mediated oncogenesis and may serve as a model to better understand factors that regulate tumor formation. SIGNIFICANCE This investigation into malignant transformation of T cells uncovers a requirement for TCR triggering, elucidates integral signaling complexes nucleated by NPM-ALK, and delineates dynamic transcriptional changes as a T cell transforms.See related commentary by Spasevska and Myklebust, p. 3160.Most primary liver cancer (PLC) cases progress mainly due to underlying chronic liver inflammation, yet the underlying mechanisms of inflammation-mediated PLC remain unclear. Here we uncover a TNF receptor II (TNFR2)-hnRNPK-YAP signaling axis in hepatic progenitor cells (HPC) essential for PLC development. TNFR2, but not TNF receptor I (TNFR1), was required for TNFα-induced activation of YAP during malignant transformation of HPCs and liver tumorigenesis. Mechanistically, heterogeneous nuclear ribonuclear protein K (hnRNPK) acted downstream of TNFα-TNFR2 signaling to directly interact with and stabilize YAP on target gene promoters genome-wide, therefore coregulating the expression of YAP target genes. Single-cell RNA sequencing confirmed the association of TNFR2-hnRNPK with YAP expression and the pathologic importance of HPC. Accordingly, expressions of TNFR2, hnRNPK, and YAP were all upregulated in PLC tissues and were strongly associated with poor prognosis of PLC including patient survival. Collectively, this study clarifies the differential roles of TNFRs in HPC-mediated tumorigenesis, uncovering a TNFR2-hnRNPK-centered mechanistic link between the TNFα-mediated inflammatory milieu and YAP activation in HPCs during PLC development. SIGNIFICANCE This work defines how hnRNPK links TNFα signaling and Hippo pathway transcription coactivator YAP in hepatic progenitor cells during primary liver tumorigenesis.Possible segregation of plasma membrane (PM) phosphoinositide metabolism in membrane lipid domains is not fully understood. We exploited two differently lipidated peptide sequences, L10 and S15, to mark liquid-ordered, cholesterol-rich (Lo) and liquid-disordered, cholesterol-poor (Ld) domains of the PM, often called raft and nonraft domains, respectively. Imaging of the fluorescent labels verified that L10 segregated into cholesterol-rich Lo phases of cooled giant plasma-membrane vesicles (GPMVs), whereas S15 and the dye FAST DiI cosegregated into cholesterol-poor Ld phases. The fluorescent protein markers were used as Förster resonance energy transfer (FRET) pairs in intact cells. An increase of homologous FRET between L10 probes showed that depleting membrane cholesterol shrank Lo domains and enlarged Ld domains, whereas a decrease of L10 FRET showed that adding more cholesterol enlarged Lo and shrank Ld Heterologous FRET signals between the lipid domain probes and phosphoinositide marker proteins suggested that phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P 2] and phosphatidylinositol 4-phosphate (PtdIns4P) are present in both Lo and Ld domains. In kinetic analysis, muscarinic-receptor-activated phospholipase C (PLC) depleted PtdIns(4,5)P 2 and PtdIns4P more rapidly and produced diacylglycerol (DAG) more rapidly in Lo than in Ld Further, PtdIns(4,5)P 2 was restored more rapidly in Lo than in Ld Thus destruction and restoration of PtdIns(4,5)P 2 are faster in Lo than in Ld This suggests that Lo is enriched with both the receptor G protein/PLC pathway and the PtdIns/PI4-kinase/PtdIns4P pathway. The significant kinetic differences of lipid depletion and restoration also mean that exchange of lipids between these domains is much slower than free diffusion predicts.