Quantitative proteomics reveals posttranslational control as a regulatory factor in primary hematopoietic stem cells.
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.
Unwin
Richard D
RD
Stem Cell and Leukaemia Proteomics Laboratory, Faculty of Medical and Human Sciences, University of Manchester, Manchester M20 4QL, UK.
Smith
Duncan L
DL
Blinco
David
D
Wilson
Claire L
CL
Miller
Crispin J
CJ
Evans
Caroline A
CA
Jaworska
Ewa
E
Baldwin
Stephen A
SA
Barnes
Kay
K
Pierce
Andrew
A
Spooncer
Elaine
E
Whetton
Anthony D
AD
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
D020543:Proteome