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Quantitative proteomics reveals post-translational control as a regulatory factor in primary hematopoietic stem cells.
Unwin, RD, Smith, D, Blinco, D, Wilson, C, Miller, C, Evans, C, Jaworska, E, Baldwin, S, Barnes, K, Pierce, A, Spooncer, E, Whetton, A
Blood. 2006;.
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Abstract
The proteome is determined by rates of transcription, translation and protein turnover. Definition of stem cell populations therefore requires a stem cell proteome signature. However, the limit to the number of primary cells available has restricted extensive proteomic analysis. We present a mass spectrometric method using an isobaric covalent modification of peptides for relative quantification (iTRAQ) which was employed to compare the proteomes of approximately 1 million long term reconstituting hematopoietic stem cells (Lin-Sca+Kit+; LSK+) and non-long term reconstituting progenitor cells (Lin-Sca+Kit-; LSK-) respectively. Extensive 2-dimensional liquid chromatography (LC) peptide separation prior to mass spectrometry (MS) enabled enhanced proteome coverage with relative quantification of 948 proteins. Of the 145 changes in the proteome 54% were not seen in the transcriptome. Hypoxia-related changes in proteins controlling metabolism and oxidative protection were observed, indicating LSK+ cells are adapted for anaerobic environments. This approach can define proteomic changes in primary samples thereby characterising the molecular signature of stem cells and their progeny.