ABSTRACT: Proteome profiles of isolated peripheral human neutrophils pretreated with purified Tamm-Horsfall protein and stimulated to undergo NETosis with PMA.
Project description:To elucidate changes in macrophage subpopulations in the presence of non-polymerizing Tamm-Horsfall Protein, we performed single cell RNAseq on mouse bone marrow derived macrophages following 18 hour treatment with either a non-polymerizing form of Tamm-Horsfall Protein or vehicle. After treatment, cells were harvested and dead cells removed prior to sorting on a 10X Chromium platform version 3 and generation of libraries. Sequencing was perfomred on an Illlumina NovaSeq 6000.
Project description:Cultured cell line UCSD-AML1 (45,XX,-7,t(3;3)(q21;q26)) and patient-derived AML194 (acute myeloid leukemia with inv3(q21;q26.2)) cells were treated in duplicate with 0 nM or 100 nM of FHD-286 for 48 hours to determine the global protein expression alterations that correlate with the cell cycle, growth inhibitory and lethal effects of treatment with a small molecule, chromatin remodeling complex protein, dual SMARCA2/SMARCA4 enzymatic activity inhibitor in MECOM rearranged AML cells with EVI1 overexpression. We also compared the global expression changes that were common between the two FHD-286-treated AML subtypes.
Project description:Patient-derived AML191 (acute myeloid leukemia with inv3(q21;q26.2)), -7) cells were treated in duplicate with 100 nM of FHD-286 for 0, 8, 16, 24 and 48 hours to determine the global protein expression alterations that correlate with the cell cycle, growth inhibitory and lethal effects of treatment with a small molecule, chromatin remodeling complex protein, dual SMARCA2/SMARCA4 enzymatic activity inhibitor in MECOM-rearranged AML cells with EVI1 overexpression.
Project description:MOLM13 cells and patient-derived de novo AML cells with MLL1 rearrangement were treated with 100 nM of FHD-286 or 100 nM of AU15330, a dual BRG1/BRM protein degrader for 48 hours. The goal was to determine the global protein expression alterations that correlate with the cell cycle, growth inhibitory and/or lethal effects of treatment with a chromatin remodeling inhibitor, FHD-286, or a BRG1/BRM protein degrader in MLL1-rearranged AML cells.
Project description:Patient-derived AML194 (acute myeloid leukemia with inv3(q21;q26.2)) cells were treated in duplicate with 0 nM or 500 nM of mivebresib, 500 nM of dual PI3K/mTOR inhibitor dactolisib or 1000 nM of IAP antagonist LCL-161 for 24 hours to determine the global protein expression alterations that correlate with the cell cycle, growth inhibitory and lethal effects of treatment with these small molecules in MECOM-rearranged AML cells with EVI1 overexpression.
Project description:We treated patient-derived mutant NPM1-and FLT3-ITD expressing AML cells with a chromatin remodeling inhibitor, FHD-286, at a dose of 100 nM, for 48 hours to determine FHD-286-mediated changes to the AML proteome.
Project description:We treated MLL1-rearranged AML MOLM13 cells with a chromatin remodeling inhibitor, FHD-286, at a dose of 100 nM, for different time intervals (up to 48 hours) to measure the time-dependent changes to the AML proteome
Project description:TcpC is a multifunctional virulence factor of uropathogenic E. coli (UPEC). Neutrophil extracellular trap formation (NETosis) is a crucial anti-infection mechanism of neutrophils. Here we show the influence of TcpC on NETosis and related mechanisms. In situ NETosis of kidneys from pyelonephritis mouse model induced by TcpC-secreting wild-type CFT073 (CFT073wt) and LPS-induced in vitro NETosis in CFT073wt- or recombinant TcpC (rTcpC)-treated neutrophils are inhibited. rTcpC enters neutrophils through caveolin-mediated endocytosis and inhibits LPS-induced production of ROS, proinflammatory cytokines and protein but not mRNA levels of peptidylarginine deiminase 4 (PAD4). rTcpC treatment enhances PAD4 ubiquitination and accumulation in proteasomes. Moreover, in vitro ubiquitination kit analyses suggest that TcpC is a PAD4-targetd E3 ubiquitin-ligase. These data suggest that TcpC inhibits NETosis primarily by serving as an E3 ligase that promotes degradation of PAD4. Our findings provide a novel mechanism underlying TcpC-mediated innate immune evasion.
Project description:Label-free quantitative proteomics was employed to compare the protein content of extracellular vesicles isolated by various differential centrifugation-based approaches from expressed prostatic secretions in urine (EPS-urine) from men with prostate cancer. The developed optimized approach improved EV purity by depleting the high-abundance urine protein Tamm-Horsfall protein (THP) and other common contaminants and achieved relative enrichment of prostate cancer-associated EV-resident proteins.