Dataset Information

Target identification for 11q13 amplicon

ABSTRACT: Proteomic strategy to define therapeutically relevant targets in cell lines that contain the 11q13 amplicon compared to those that do not and to ascertain which genes are amplified at the protein level and, concomitantly, are key drivers for tumor growth and/or maintenance. Furthermore, so called passenger genes that are amplified with driver genes and a manifest on the cell surface can be attractive targets for an antibody – drug conjugate approach (ADC).

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Permanent Cell Line Cell, Cell Culture

DISEASE(S): Neck Carcinoma

SUBMITTER: Heather Hoover

LAB HEAD: Nancy Finkel

PROVIDER: PXD002486 | Pride | 2015-08-04

REPOSITORIES: Pride

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Publications

Quantitative Proteomic Verification of Membrane Proteins as Potential Therapeutic Targets Located in the 11q13 Amplicon in Cancers.

Hoover Heather H Li Jun J Marchese Jason J Rothwell Christopher C Borawoski Jason J Jeffery Douglas A DA Gaither L Alex LA Finkel Nancy N

Journal of proteome research 20150729 9

Tumor types can be defined cytologically by their regions of chromosomal amplification, which often results in the high expression of both mRNA and proteins of certain genes contained within the amplicon. An important strategy for defining therapeutically relevant targets in these situations is to ascertain which genes are amplified at the protein level and, concomitantly, are key drivers for tumor growth or maintenance. Furthermore, so-called passenger genes that are amplified with driver genes ...[more]

PMID: 26151158

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Project description:In the past years several protocols for the proteomic profiling of plasma membrane proteins have been described. Nevertheless, comparative analyses have mainly focused on different variations of one approach [1-3]. To pave the way to high-performance differential plasma membrane proteomics, we compared sulfo-NHS-SS-biotinylation, aminooxy-biotinylation and surface coating with silica beads to isolate plasma membrane proteins for subsequent analysis by one-dimensional gel-free liquid chromatography mass spectrometry. Absolute and relative numbers of plasma membrane proteins and reproducibility parameters on a qualitative and quantitative level were assessed. Sulfo-NHS-SS-biotinylation outperformed aminooxy-biotinylation and surface coating using silica beads for most of the monitored criteria. We further simplified this procedure by introducing a competitive biotin elution strategy, for which the average plasma membrane annotated protein fraction amounted to 54 % (347 proteins). Moreover, purified non-plasma membrane annotated proteins and plasma membrane annotated proteins displayed similarly high reproducibility suggesting specific co-purification. In fact, the non-plasma membrane annotated data was extremely enriched for direct interactors of purified plasma membrane proteins. Computational analysis using additional databases and prediction tools revealed that in total over 90 % of the purified proteins were associated with the plasma membrane. The modified sulfo-NHS-SS-biotinylation protocol was validated by tracking changes in the plasma membrane proteome composition induced by genetic alteration and drug treatment. GPI-anchored proteins were depleted in plasma membrane purifications from cells deficient in the GPI transamidase component PIGS; and treatment of cells with tunicamycin significantly reduced the abundance of N-glycoproteins in surface purifications. Altogether, this study introduces an improved, filter-free sulfo-NHS-SS-biotinylation protocol and demonstrates it to be a specific, effective and reproducible method to isolate proteins associated with the plasma membrane, thus enabling future large-scale comparative cell surface mappings.

Dataset Information

Target identification for 11q13 amplicon

Publications

Quantitative Proteomic Verification of Membrane Proteins as Potential Therapeutic Targets Located in the 11q13 Amplicon in Cancers.

Similar Datasets

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