Ifitm3 was previously identified as an endosomal protein that blocks viral infection 1 – 3 . Studying clinical cohorts of B-cell leukemia and lymphoma patients, we identified IFITM3 as a strong predictor of poor outcome. In normal resting B-cells, Ifitm3 was minimally expressed and mainly localized in endosomes. However, B-cell receptor (BCR) engagement induced expression of Ifitm3 and phosphorylation at Y20, resulting in accumulation at the cell surface. In B-cell leukemia, oncogenic kinases phosphorylate IFITM3-Y20, causing constitutive plasma membrane localization. Ifitm3 ˉ / ˉ naïve B-cells developed at normal numbers; however, germinal center formation and production of antigen-specific antibodies were compromised. Oncogenes that induce development of leukemia and lymphoma failed to transform Ifitm3 ˉ / ˉ B-cells. Conversely, the phospho-mimetic IFITM3-Y20E induced oncogenic PI3K-signaling and initiated transformation of pre-malignant B-cells. Mechanistic experiments revealed that Ifitm3 functions as PIP3-scaffold and central amplifier of PI3K signaling. PI3K signal-amplification depends on Ifitm3 scaffolding PIP3-accumulation via two lysine residues (K83 and K104) in its conserved intracellular loop. In Ifitm3 ˉ / ˉ B-cells, lipid rafts were depleted of PIP3, resulting in defective expression of >60 lipid raft-associated surface receptors, impaired BCR-signaling and cellular adhesion. We conclude that phosphorylation of IFITM3 upon B-cell antigen-encounter induces a dynamic switch from antiviral effector functions in endosomes to a PI3K-amplification loop at the cell surface. IFITM3-dependent amplification of PI3K-signaling in part downstream of the BCR is critical to enable rapid expansion of B-cells with high affinity to antigen. In addition, multiple oncogenes depend on IFITM3 to assemble PIP3-dependent signaling complexes and amplify PI3K-signaling for malignant transformation.

Genotyping graft livers by short tandem repeats after human living-donor liver transplantation (n ¼ 20) revealed the presence of recipient or chimeric genotype cases in hepatocytes (6 of 17, 35.3%), sinusoidal cells (18 of 18, 100%), cholangiocytes (15 of 17, 88.2%) and cells in the periportal areas (7 of 8, 87.5%), suggesting extrahepatic cell involvement in liver regeneration. Regarding extrahepatic origin, bone marrow mesenchymal stem cells (BM-MSCs) have been suggested to contribute to liver regeneration but compose a heterogeneous population. We focused on a more specific subpopulation (1-2% of BM-MSCs), called multilineage-differentiating stress-enduring (Muse) cells, for their ability to differentiate into liver-lineage cells and repair tissue. We generated a physical partial hepatectomy model in immunodeficient mice and injected green fluorescent protein (GFP)-labeled human BM-MSC Muse cells intravenously (n ¼ 20). Immunohistochemistry, fluorescence in situ hybridization and species-specific polymerase chain reaction revealed that they integrated into regenerating areas and expressed liver progenitor markers during the early phase and then differentiated spontaneously into major liver components, including hepatocytes (%74.3% of GFP-positive integrated Muse cells), cholangiocytes (%17.7%), sinusoidal endothelial cells (%2.0%), and Kupffer cells (%6.0%). In contrast, the remaining cells in the BM-MSCs were not detected in the liver for up to 4 weeks. These results suggest that Muse cells are the predominant population of BMMSCs that are capable of replacing major liver components during liver regeneration.

Cancer relapse occurs with substantial frequency even after treatment with curative intent. Here we studied drug-tolerant colonies (DTCs), which are subpopulations of cancer cells that survive in the presence of drugs. Proteomic characterization of DTCs identified stemness- and epithelial-dominant subpopulations, but functional screening suggested that DTC formation was regulated at the transcriptional level independent from protein expression patterns. We consistently found that α-amanitin, an RNA polymerase II (RNAPII) inhibitor, effectively inhibited DTCs by suppressing TAF15 expression, which binds to RNA to modulate transcription and RNA processing. Sequential administration of α-amanitin and cisplatin extended overall survival in a cancer-relapse mouse model, namely peritonitis carcinomatosa. Therefore, post-treatment cancer relapse may occur through non-distinct subpopulations and may be effectively prevented by α-amanitin to disrupt transcriptional machinery, including TAF15.

Malignant transformation typically involves multiple genetic lesions whose combined activity gives rise to cancer 1 . Our analysis of 1,148 patient-derived B-cell leukemia (B-ALL) samples revealed that individual mutations did not promote leukemogenesis unless they converged on one single oncogenic pathway characteristic for the differentiation stage of transformed B cells. Mutations not aligned with the central oncogenic driver activated divergent pathways and subverted transformation. Oncogenic lesions in B-ALL frequently mimic cytokine receptor signaling at the pro-B cell stage (through activation of STAT5) 2 – 4 or the pre-B cell receptor in more mature cells (through activation of ERK) 5 – 8 . STAT5- and ERK-activating lesions were frequently found but only co-occurred in ~3% of cases ( P =2.2E-16). Single-cell mutation and phosphoprotein analyses revealed the segregation of oncogenic STAT5- or ERK-activation to competing clones. STAT5 and ERK engaged opposing biochemical and transcriptional programs orchestrated by MYC and BCL6, respectively. Genetic reactivation of the divergent (suppressed) pathway came at the expense of the principal oncogenic driver and reversed transformation. Conversely, deletion of divergent pathway components accelerated leukemogenesis. Thus, persistence of divergent signaling pathways represents a powerful barrier to transformation while convergence on one principal driver defines a central event in leukemia-initiation. Pharmacological reactivation of suppressed divergent circuits strongly synergized with inhibition of the principal oncogenic driver. Hence, reactivation of divergent pathways can be leveraged as a previously unrecognized strategy to deepen treatment responses.

Despite of remarkable improvement of postoperative 5-FU–based adjuvant chemotherapy, the relapse rate of gastric cancer patients who undergo curative resection followed by the adjuvant chemotherapy remains substantial. Therefore, it is important to identify prediction markers for the chemotherapeutic efficacy of 5-FU. We recently identified NF-κB as a candidate relapse prediction biomarker in gastric cancer. To evaluate the biological significance of NF-κB in the context of 5-FU–based chemotherapy, we analyzed the NF-κB-dependent biological response upon 5-FU treatment in gastric cancer cell lines. Seven genes induced by 5-FU treatment in an NF-κB-dependent manner were identified, five of which are known p53 targets. Knockdown of RELA, which encodes the p65 subunit of NF-κB, decreased both p53 and p53 target protein levels. In contrast, NF-κB was not affected by TP53 knockdown. We also demonstrated that cell lines bearing Pro/Pro homozygosity in codon72 of p53 exon4, which is important for NF-κB binding to p53, are more resistant to 5-FU than those with Arg/Arg homozygosity. We conclude that NF-κB plays an important role in the response to 5-FU treatment in gastric cancer cell lines, with a possible compensatory function of p53. These results suggest that NF-κB is a potential 5-FU-chemosensitivity prediction marker that may reflect 5-FU-induced stress-response pathways, including p53.