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Blood, 15 June 2006, Vol. 107, No. 12, pp. 4687-4694.
Prepublished online as a Blood First Edition Paper on February 28, 2006; DOI 10.1182/blood-2005-12-4995.
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HEMATOPOIESIS
Quantitative proteomics reveals posttranslational control as a regulatory factor in primary hematopoietic stem cells
Richard D. Unwin,
Duncan L. Smith,
David Blinco,
Claire L. Wilson,
Crispin J. Miller,
Caroline A. Evans,
Ewa Jaworska,
Stephen A. Baldwin,
Kay Barnes,
Andrew Pierce,
Elaine Spooncer, and
Anthony D. Whetton
From the Stem Cell and Leukaemia Proteomics Laboratory, Faculty of Medical and Human Sciences, University of Manchester, Kinnaird House, Manchester; the Bioinformatics Group, Paterson Institute for Cancer Research, Christie Hospital, Manchester; and the Institute of Membrane and Systems Biology, Faculty of Biological Sciences, University of Leeds, United Kingdom.
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 that LSK+ cells are adapted for anaerobic environments. This approach can define proteomic changes in primary samples, thereby characterizing the molecular signature of stem cells and their progeny.
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