Project PXD000185

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Summary

Title

Kinase Substrate Enrichment Analysis Provides Insights into the Heterogeneity of Signaling Pathway Activation in Leukemia Cells

Description

Experiments were performed in AML (Acute Myeloid Leukemia) cell lines and in primary cells coming from patients with AML and healthy donors. The AML cell lines P31/Fuj and Kasumi-1 were treated with vehicle (DMSO), 100 nM or 1000 nM of the PI3K/mTOR inhibitor AZ123 or the mTOR inhibitor Ku0063794 for 2h. Both cell lines were treated also with vehicle and 1000 nM of the PI3K/mTOR inhibitor PI103. 6 replicates per condition were done. AML primary cells and GMPBs (G-CSF-mobilized peripheral blood cell) from patients and healthy donors respectively were treated or untreated with 100 mM sodium pervanadate for 30 min. Samples 45-46 correspond to the GMPBs and the rest correspond to AML patients. All samples were digested with trypsin and subjected to phosphoenrichment usin TiO2. Phosphopeptides were run in a LTQ-Orbitrap-XL. Peaks lists were generated with Mascot Distiller (version 2.3) in MGF format and Database searches were with Mascot Server (version 2.3) against the SwissProt database restricted to human sequences (release December 2011) and trypsin cleavage. Restrictions were 7ppm for parent ions and 600 mmu for fragment masses. Allowed modifications were phosphorylation of Ser/Thr/Tyr, pyro-Glu (N-term) and methionine oxidation and one miss-cleavage allowed. Quantification was by label-free using peak heights of extracted ion chromatograms (XICs) constructed with narrow mass windows (7ppm) and time windows (1.5 minutes). Pescal, a computer program written in house, was used to automate the generation of XICs and to calculate peak heights.

Sample Processing Protocol

See details in reference(s) : 23532336

Data Processing Protocol

See details in reference(s) : 23532336

Contact

Pedro Casado-Izquierdo, Cell Signalling

Submission Date

22/03/2013

Publication Date

09/04/2013

Publication

    Casado P, Rodriguez-Prados JC, Cosulich SC, Guichard S, Vanhaesebroeck B, Joel S, Cutillas PR; Kinase-substrate enrichment analysis provides insights into the heterogeneity of signaling pathway activation in leukemia cells., Sci Signal, 2013 Mar 26, 6, 268, rs6, PubMed: 23532336

Assay

Page 1 2
Page size 10
Showing 1 - 10 of 17 results
# Accession Title Proteins Peptides Unique Peptides Spectra Identified Spectra View in Reactome
1 28859 Effects of AZ123 in Kas Cells File 1 of 2 1043 8320 1891 41378 8320
2 28860 Effects of AZ123 in Kas Cells File 2 of 2 436 2932 625 38390 2932
3 28861 Effects of AZ123 in P31 Cells File 1 of 2 969 8086 1643 37268 8086
4 28862 Effects of AZ123 in P31 Cells File 2 of 2 681 5347 1073 43697 5347
5 28863 Effects of Ku in Kas Cells File 1 of 2 711 5728 1109 45325 5728
6 28864 Effects of Ku in Kas Cells File 2 of 2 794 7279 1321 41643 7279
7 28865 Effects of Ku in P31 Cells File 1 of 2 525 2861 801 50158 2861
8 28866 Effects of Ku in P31 Cells File 2 of 2 757 5206 1231 47794 5206
9 28867 Effects of PI103 in Kas Cells File 1 of 2 511 2872 775 13681 2872
10 28868 Effects of PI103 in Kas Cells File 2 of 2 336 1811 478 11847 1811