title: Quantitative proteomics reveals posttranslational control as a regulatory factor in primary hematopoietic stem cells.
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 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.
authors.affiliation: Stem Cell and Leukaemia Proteomics Laboratory, Faculty of Medical and Human Sciences, University of Manchester, Manchester M20 4QL, UK.
authors.forename: Richard D
authors.initials: RD
authors.lastname: Unwin
authors.forename: Duncan L
authors.initials: DL
authors.lastname: Smith
authors.forename: David
authors.initials: D
authors.lastname: Blinco
authors.forename: Claire L
authors.initials: CL
authors.lastname: Wilson
authors.forename: Crispin J
authors.initials: CJ
authors.lastname: Miller
authors.forename: Caroline A
authors.initials: CA
authors.lastname: Evans
authors.forename: Ewa
authors.initials: E
authors.lastname: Jaworska
authors.forename: Stephen A
authors.initials: SA
authors.lastname: Baldwin
authors.forename: Kay
authors.initials: K
authors.lastname: Barnes
authors.forename: Andrew
authors.initials: A
authors.lastname: Pierce
authors.forename: Elaine
authors.initials: E
authors.lastname: Spooncer
authors.forename: Anthony D
authors.initials: AD
authors.lastname: Whetton
mesh_terms: D000818:Animals; D015687:Cell Hypoxia; D002853:Chromatography, Liquid; D006412:Hematopoietic Stem Cells / Q000502:physiology*; D013058:Mass Spectrometry; D051379:Mice; D010084:Oxidation-Reduction; D011499:Protein Processing, Post-Translational / Q000502:physiology*; D020543:Proteome / Q000235:genetics / Q000378:metabolism*; D040901:Proteomics
chemical_list: D020543:Proteome