Summary of Study ST002412
This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001552. The data can be accessed directly via it's Project DOI: 10.21228/M8342Z This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.
| Study ID | ST002412 |
| Study Title | Metabolic effects of the protein kinase R |
| Study Type | Biomedical research |
| Study Summary | Spleen-derived macrophage from WT or Eif2ak2-/- (gene encoding PKR protein kinase) mice are treated with a synthetic RNA mimetic (polyinosinic:polycytidylic acid) to activate the kinase and metabolites were collected for analysis. The data identified 325 putative metabolites in the cell extracts, with a large number of significant differences between the Eif2ak2- /- and WT sample groups. Metabolite levels are predominantly suppressed in the WT compared to the Eif2ak2-/- cells, with depletion of specific metabolites in amino acid, carbohydrate, lipid and nucleotide pathways, while several amino acid metabolites were significantly elevated in the WT cells compared to the Eif2ak2-/-. The changes appear to delineate a pseudo-starvation response in the WT cells. Phosphate energy metabolism is altered with decreased creatine and phosphocreatine and a compensatory increase in phosphorylated guanidinoacetate in the WT compared to the Eif2ak2- /- cells. There appears to be a constraint in glycolysis in the WT cells, most clearly in the pentose phosphate pathway. |
| Institute | Hudson Institute of Medical Research |
| Department | CIIID |
| Laboratory | Molecular Immunology |
| Last Name | Sadler |
| First Name | Anthony |
| Address | 27-31 Wright st, Clayton, VIC 3168 |
| Anthony.sadler@hudson.org.au | |
| Phone | +61 4 85722722 |
| Submit Date | 2022-12-15 |
| Num Groups | 2 |
| Total Subjects | NA |
| Num Males | NA |
| Num Females | NA |
| Study Comments | KO vs WT |
| Publications | Suppression of the nucleic acid precursor ribose 5-phosphate by RNA-mediated antiviral immunity |
| Raw Data Available | Yes |
| Raw Data File Type(s) | raw(Thermo) |
| Analysis Type Detail | LC-MS |
| Release Date | 2023-01-04 |
| Release Version | 1 |
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Collection:
| Collection ID: | CO002494 |
| Collection Summary: | Exogenous genes were stably expressed and endogenous gene expression suppressed in murine macrophages by transduction with lentiviral constructs. Gene open reading frames were cloned as NheI-BamHI (New England Biolabs) fragments in the plentiCRISPR v2-Blast (Mohan Babu, Addgene plasmid # 83480; http://n2t.net/addgene:83480; RRID: Addgene_83480). Knockdown of endogenous transcripts was achieved with short hairpin RNA cloned into pLKO.1-puro. Lentiviral particles were packaged using a Lenti-X mix (Takara) by transfection of the plasmid components into HEK293FT cells with Lipofectamine 2000 (Invitrogen). The culture supernatant was collected after 48 hours, centrifuged at 10 000 g, then pipetted onto adherent macrophages. Transduced cells were isolated over 2-3 month by increasing selection (from 1 to 20 ug/mL) with puromycin (Thermo Fisher Scientific) and/or blasticidin (InvivoGen), for pLKO-1-puro-shRNA and plentiCRISPR v2-blast, respectively. Recombinant cells were maintained with antibiotics in Dulbecco’s modified Eagle’s Medium (DMEM, GibCo) supplemented with 10% foetal bovine serum (Bovogen) in a humidified 5% CO 2 incubator at 37 °C. Growth on different carbon sources was tested by plating 6x10 3 cells per well of a 24 well plate with glucose free DMEM (ThermoFisher Scientific) supplemented with 10% FBS (not dialysed), 20 ug/mL puromycin and 20 uM of each sugar. After 72 hours the cells were lifted with trypsin and counted by automation (Countess, Invitrogen). |
| Sample Type: | Macrophages |
