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PXD012573 is an original dataset announced via ProteomeXchange.

Dataset Summary
TitlePhosphoproteomics analysis of Erk signaling in embryonic stem cell differentiation
DescriptionMass spectrometry-based proteomics was applied to unravel Erk signaling in mouse embryonic stem cells (ESCs). A previously described engineered ESCs system for Erk induction via a drug-inducible cRAF-ErT2 fusion (Hamilton, W. B. & Brickman, J. M., Cell Rep 2014) was used in two different setups measuring the phosphoproteome in an 11-plex tandem mass tag (TMT)-based approach as described below. Fgf4 stimulation and MEK inhibition: ESCs were stimulated with Fgf4 (40 nM, recombinant, mouse, 5 minutes) following pretreatment (30 minutes) with either DMSO (1:10000) or MEK inhibition (MEKi: 1 microM PD0325901): The experimental treatment conditions and TMT11-plex labeling are specified below: 126: DMSO, unstimulated control replicate 1 127N: DMSO, unstimulated control replicate 2 127C: DMSO, unstimulated control replicate 3 128N: DMSO + Fgf4 replicate 1 128C: DMSO + Fgf4 replicate 2 129N: DMSO + Fgf4 replicate 3 129C: MEKi + Fgf4 replicate 1 130N: MEKi + Fgf4 replicate 2 130C: MEKi + Fgf4 replicate 3 131N: MEKi replicate 1 131C: MEKi replicate 2 Analog sensitive Erk2 setup: In the ESC system the ERK2-/- ESCs (Hamilton et al., PloS ONE, 2013) were engineered to stably express an inducible cRAF-ErT2 fusion protein expressed from the same transgene as a FLAG-tagged analogue sensitive Erk2Q103A. Erk1 was subsequently removed by CRISPR-Cas9 mutagenesis using sgRNAs flanking exon 2. Cells were pre-treated for 30 minutes with either the ATP-mimetic compound 1-NM-PP1 (2 microM) (Endo et al., Biochem Biophys Res Commun 2006), the Mek1 inhibitor PD0325901 (1 microM), or DMSO (0.01%), followed by induction or not of the cRAF-ErT2 fusion by (Z)-4-Hydroxytamoxifen (4OHT) (250 nM) for 2 hours as indicated. The experimental treatment conditions and TMT11-plex labeling are specified below: 126: DMSO pretreatment, non-induced control, replicate 1 127N: DMSO pretreatment, non-induced control, replicate 2 127C: DMSO pretreatment, non-induced control, replicate 3 128N: DMSO pretreatment, 4OHT-induction, replicate 1 128C: DMSO pretreatment, 4OHT-induction, replicate 2 129N: DMSO pretreatment, 4OHT-induction, replicate 3 129C: PP1 pretreatment, 4OHT-induction, replicate 1 130N: PP1 pretreatment, 4OHT-induction, replicate 2 130C: PP1 pretreatment, 4OHT-induction, replicate 3 131N: MEKi pretreatment, 4OHT-induction, replicate 1 131C: MEKi pretreatment, 4OHT-induction, replicate 2
ReviewLevelPeer-reviewed dataset
DatasetOriginOriginal dataset
RepositorySupportUnsupported dataset by repository
PrimarySubmitterKristina Bennet Emdal
SpeciesList scientific name: Mus musculus (Mouse); NCBI TaxID: 10090;
ModificationListphosphorylated residue
InstrumentQ Exactive
Dataset History
RevisionDatetimeStatusChangeLog Entry
02019-01-31 01:36:31ID requested
12019-08-12 01:43:33announced
22019-11-18 01:03:47announced2019-11-18: Updated publication reference for PubMed record(s): 31695196.
Publication List
Hamilton WB, Mosesson Y, Monteiro RS, Emdal KB, Knudsen TE, Francavilla C, Barkai N, Olsen JV, Brickman JM, Dynamic lineage priming is driven via direct enhancer regulation by ERK. Nature, 575(7782):355-360(2019) [pubmed]
Keyword List
curator keyword: Biological
submitter keyword: Mouse, embryonic stem cells, LC-MSMS
Contact List
Jesper Velgaard Olsen
contact affiliationNovo Nordisk Foundation Center for Protein Research, Faculty of Health & Medical Sciences, University of Copenhagen, Blegdamsvej 3B, DK-2200 Copenhagen, Denmark
contact emailjesper.olsen@cpr.ku.dk
lab head
Kristina Bennet Emdal
contact affiliationNNF Center for Protein Research, Proteomics Program, Faculty of Health and Medical Sciences, University of Copenhagen.
contact emailkristina.emdal@cpr.ku.dk
dataset submitter
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