Proteomics

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Protrusion vs Cell-body SILAC proteomics

ABSTRACT: In mesenchymal-like cell migration, cells need to polarise into a protrusive front and a retracting cell body. To understand this process better, we set out to quantify the distribution of cellular proteins between protrusions and cell-body by proteomics, using MDA-MB-231 cells, a highly invasive breast cancer cell-line. We utilised a transwell filter based fractionation method in conjugation with SILAC proteomics. In this method, cells are seeded on top of 3μm transwell filters to enable protrusions to form through the pores of the filters but to prevent the cell-bodies passing through due to the small size of the pores, thus resulting in separation of protrusions and cell-bodies on opposite sides of the filter, which can then be lysed and prepared separately. Prepration of protrusion and cell-body fractions from heavy and light SILAC labelled cells then allows for reciprocal mixing and quantification of proteins between protrusions and cell-body. In this study, we determined the relative distribution of 3240 proteins between protrusions and cell-body from two SILAC mixes. Associated ArrayExpress data: E-MTAB-2546.

OTHER RELATED OMICS DATASETS IN: PXD000914PXD002649

INSTRUMENT(S): LTQ Orbitrap Velos

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

SUBMITTER: Dr Christopher J. Marshall 

PROVIDER: MSV000079570 | MassIVE | Thu Mar 10 12:38:00 GMT 2016

SECONDARY ACCESSION(S): PXD000914

REPOSITORIES: MassIVE

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Publications

Global Analysis of mRNA, Translation, and Protein Localization: Local Translation Is a Key Regulator of Cell Protrusions.

Mardakheh Faraz K FK   Paul Angela A   Kümper Sandra S   Sadok Amine A   Paterson Hugh H   Mccarthy Afshan A   Yuan Yinyin Y   Marshall Christopher J CJ  

Developmental cell 20151101 3

Polarization of cells into a protrusive front and a retracting cell body is the hallmark of mesenchymal-like cell migration. Many mRNAs are localized to protrusions, but it is unclear to what degree mRNA localization contributes toward protrusion formation. We performed global quantitative analysis of the distributions of mRNAs, proteins, and translation rates between protrusions and the cell body by RNA sequencing (RNA-seq) and quantitative proteomics. Our results reveal local translation as a  ...[more]