Project description:Transcriptional profiling of Murine BaF3 cells infected with MPLW515L grown under either normal conditions (Naive) or in 0.8 uM INCB18424 for 4-6 weeks (Persistent). Naive and Persistent cells were then treated with either DMSO (Control) or 0.8 uM INCB18424 for 4 hours. Goal was to determine transcriptional changes conditioned upon sensitivity/resistance of BaF3 MPLW515L mutants to JAK1/2 specific inhibitor.
Project description:Transcriptional profiling of Murine BaF3 cells infected with MPLW515L grown under either normal conditions (Naive) or in 0.8 uM INCB18424 for 4-6 weeks (Persistent). Naive and Persistent cells were then treated with either DMSO (Control) or 0.8 uM INCB18424 for 4 hours. Goal was to determine transcriptional changes conditioned upon sensitivity/resistance of BaF3 MPLW515L mutants to JAK1/2 specific inhibitor. 3 condition experiment consiting of: 1) Naive cells treated with DMSO (Control) , 2) Naïve cells treated with 0.8 uM INCB18424 for 4 hours (Acute) and 3) Persistent cells treated with 0.8 uM INCB18424 for 4 hours (Persistent). Biological replicates: 3 DMSO control replicates, 3 Acute replicates, 3 Persistent replicates.
Project description:The activation of PD-1 (Programmed Death receptor-1) on T cells can cause T cell exhaustion and immune tolerance. Some tumors up-regulate the expression of the ligand of PD-1, namely PD-L1 (Programmed Death Receptor-Ligand 1), thus preventing anti-tumor immune response and promoting immune-escape. Previous studies have shown that JAK2 (Janus Kinase 2) signaling can promote PD-L1 expression in Hodgkin Lymphoma. In Myeloproliferative Neoplasms (MPN), JAK2 is frequently characterized by the the presence of the point-mutation V617F, which leads to its constitutive activation and to uncontrolled cell proliferation and survival. Accordingly, tumor cell lines expressing JAK2 V617F express higher levels of PD-L1 as compared to tumor cell lines negative for such mutations. In this experiment, we transfected BaF3 cells with a vector (plasmid for Murine Stem Cell Virus) containing the gene for JAK2 with the point-mutation V617F. As control, we used BaF3 cells transfected with the same vector, but without the gene for JAK2 V617F (empty vector). Both the cell lines (with/without JAK2 V617F) were co-cultured with primary murine T cells. When co-cultured with BaF3 cells expressing JAK2 V617F, T cells upregulated genes connected to senescence pathways, showed increased apoptosis, less cytokine production, and displayed other forms of dysfunction which can be associated with the activation of PD-1.
Project description:The importance of unanchored Ub in innate immunity has been shown only for a limited number of unanchored Ub-interactors. We investigated what additional cellular factors interact with unanchored Ub and whether unanchored Ub plays a broader role in innate immunity. To identify unanchored Ub-interacting factors from murine lungs, we used His-tagged recombinant poly-Ub chains as bait. These chains were mixed with lung tissue lysates and protein complexes were isolated with Ni-NTA beads. Sample elutions were subjected to mass spectrometry (LC-MSMS) analysis.
