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Chikungunya virus glycoprotein targeting of host factors increases viral fitness in human macrophage

ABSTRACT: Proteomic analysis of Chikungunya virus E1 and E2 interacting proteins during macrophage infection

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Homo Sapiens (ncbitaxon:9606)

SUBMITTER: James Wohlschlegel

PROVIDER: MSV000094494 | MassIVE | Mon Apr 08 14:58:00 BST 2024

REPOSITORIES: MassIVE

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Inhibition of Ubiquitin Thioester Transfer by Targeting a Helix-in-Groove Interaction between E1 and E2

Project description:In eukaryotes, E1 initiates the ubiquitin cascade by adenylation and thioesterification of the ubiquitin C-terminus and subsequent transfer of ubiquitin to E2 enzymes. A clinical-grade small molecule that binds to the E1 ATP binding site and covalently derivatizes the ubiquitin C-terminus effectively shuts down E1 enzymatic activity. However, mutation at or near the ATP binding site of E1 causes resistance, mandating alternative approaches to blocking what is otherwise a promising cancer target. Here, we identified a helix-in-groove interaction between the N-terminal alpha-1 helix of E2 and a pocket within the ubiquitin fold domain of E1 as a druggable site of protein interaction. By generating and optimizing stapled alpha-helical peptides (SAHs) modeled after the E2 alpha-1 helix, we achieve site-specific engagement of E1, induce a consequential conformational change, and effectively block E1 enzymatic activity, resulting in a generalized disruption of E2 ubiquitin-charging that suppresses ubiquitination of cellular proteins. Thus, we provide a blueprint for an alternative E1-targeting strategy for the treatment of cancer. Hydrogen exchange mass spectrometry was used to characterize the predominant E1 enzyme in mammals (UBE1, a 118 kDa multi-domain enzyme that catalyzes both ubiquitin adenylation and thioesterification) in the unbound state. We then interrogated the structural impact of UBE1 interaction with the stapled peptide SAH-UBE2A and several mutants. The observed peptide-induced exposure of the ubiquitin-fold domain (UFD) linker hinge in UBE1 was consistent with an inhibitory mechanism whereby SAH-UBE2A locks UBE1 into its proximal UFD conformation.

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The major pre- and post-menopausal estrogens play opposing roles in obesity driven mammary inflammation and breast cancer development

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Expression data from murine bone marrow erythroid progenitor cells at two early stages of development.

2012-11-17 | GSE42346 | GEO

Expression data from murine bone marrow erythroid progenitor cells at two early stages of development.

Project description:This study was designed to define erythropoietin (EPO) regulated genes in murine bone marrow erythroid progenitor cells at two stages of development, designated E1, and E2. E1 cells correspond to CFUe- like progenitors, while E2 cells are proerythroblasts. 14 samples were analyzed. 8 samples belong to the E1 stage and the remaning 6 samples correspond to the E2 stage. In the E1 stage, there are 4 controls and 4 treatment samples. The 4 samples in each group are biological replicates. Likewise in the E2 stage, there are 3 control and 3 treatment samples and the 3 samples in each group are biological replicates.

2012-11-17 | E-GEOD-42346 | biostudies-arrayexpress

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Project description:Primary objectives: Characterization of the macrophage population subset that is modulated by enteric neurons Primary endpoints: Characterization of the macrophage population subset that is modulated by enteric neurons via RNA sequencing

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Embryo Mortality with an Impaired Interferon-Tau Release Mounts a Massive T-Helper Immune Response in Lactating Holstein-Friesian cows

Project description:Embryo mortality (EM) contributes to infertility, but its exact mechanism is poorly understood. It was hypothesized that bovine EM pregnancies have impaired conceptus-derived interferon-tau (IFNT) release and action and are associated with altered transcriptome responses. The objective was to discover transcriptome response in EM tissues (endometrium [ENDO], corpus luteum [CL] and peripheral blood mononuclear cells [PBMC]) in lactating Holstein-Friesian cows. Two experiments (E1, E2) in day 16 pregnant (exposed to semen; E1:n=13, E2:n=15) or non-pregnant (NP; not exposed to semen; E1:n=7, E2:n=7) cows were completed. Uterine flushings (UF) and tissues were collected. Pregnant cows were also classified based on conceptus morphology and appearance for EM (E1:n=5, E2:n=6) or normal (N) conceptuses (E1:n=8, E2:n=9). Conceptuses and tissues were RNA sequenced and analyzed. The N conceptuses were longer (P<0.05) than EM conceptuses. The IFNT protein concentrations in UF were greater in N compared to EM and NP cows in E1 but not for E2. The ENDO conjugated ISG15 concentrations were greater in N (E1) than EM and NP cows but N (E2) were only greater than NP not EM cows. The major up-regulated canonical pathway in EM conceptuses was T helper 1 (Th1) and Th2. The ENDO had a massive increase in interferon stimulated genes in N and EM compared to NP cows. Estradiol-associated genes were up-regulated in EM compared to N ENDO for E2. The PBMC in E1 reflected up-regulation of genes associated Th1 immune responses in EM compared to N cows. Luteolytic pathways were upregulated in EM CL compared to N cows. This disruption of maternal recognition of pregnancy in EM pregnancies entails a massive T helper immune response within the conceptus, estradiol re-modelling of the ENDO, abnormal immune system in PBMC, luteolysis cascade in CL and, potentially, loss of pregnancy.

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Project description:In this project, the protein mixture in the adhesive strip samples was analyzed by means of intragel enzymatic hydrolysis and high-resolution biomass spectrometry. Among them, 10 ubiquitination modification sites were identified in the samples E1+E2+HERC5+ISG15+N, the target protein was identified in the samples E1+E2+HERC5+N and N, but the target modification was not identified on the target protein.

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E2-RING expansion of the NEDD8 cascade confers specificity to cullin modification

Project description:Ubiquitin and ubiquitin-like proteins (UBLs) are directed to targets by cascades of E1, E2, and E3 enzymes. The largest ubiquitin E3 subclass consists of cullin-RING ligases (CRLs), which contain one each of several cullins (CUL1, -2, -3, -4, or -5) and RING proteins (RBX1 or -2). CRLs are activated by ligation of the UBL NEDD8 to a conserved cullin lysine. How is cullin NEDD8ylation specificity established? Here we report that, like UBE2M (also known as UBC12), the previously uncharacterized E2 UBE2F is a NEDD8-conjugating enzyme in vitro and in vivo. Biochemical and structural analyses indicate how plasticity of hydrophobic E1-E2 interactions and E1 conformational flexibility allow one E1 to charge multiple E2s. The E2s have distinct functions, with UBE2M/RBX1 and UBE2F/RBX2 displaying different target cullin specificities. Together, these studies reveal the molecular basis for and functional importance of hierarchical expansion of the NEDD8 conjugation system in establishing selective CRL activation. Mol Cell 33:483-495, 2009. Keywords: Comparison of gene expression profiles

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Macrophage

Project description:Macrophage

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EPO-regulated targets in E1, E2, and E3 erythroid progenitors

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