Identification of cell surface proteins for antibody-based selection of human embryonic stem cell-derived cardiomyocytes.
The absence of identified cell surface proteins and corresponding antibodies to most differentiated derivatives of human embryonic stem cells (hESCs) has largely limited selection of specific cell types from mixed cell populations to genetic approaches. Here, we describe the use of mass spectrometry (MS)-based proteomics on cell membrane proteins isolated from hESCs that were differentiated into cardiomyocytes to identify candidate proteins for this particular lineage. Quantitative MS distinguished cardiomyocyte-specific plasma membrane proteins that were highly enriched or detected only in cardiomyocytes derived from hESCs and human fetal hearts compared with a heterogeneous pool of hESC-derived differentiated cells. For several candidates, cardiomyocyte-specific expression and cell surface localization were verified by conventional antibody-based methodologies. Using an antibody against elastin microfibril interfacer 2 (EMILIN2), we demonstrate that cardiomyocytes can be sorted from live cell populations. Besides showing that MS-based membrane proteomics is a powerful tool to identify candidate proteins that allow purification of specific cell lineages from heterogeneous populations, this approach generated a plasma membrane proteome profile suggesting signaling pathways that control cell behavior.
Van Hoof
Dennis
D
Hubrecht Institute, Developmental Biology and Stem Cell Research, Uppsalalaan 8, 3584 CT Utrecht, The Netherlands.
Dormeyer
Wilma
W
Braam
Stefan R
SR
Passier
Robert
R
Monshouwer-Kloots
Jantine
J
Ward-van Oostwaard
Dorien
D
Heck
Albert J R
AJ
Krijgsveld
Jeroen
J
Mummery
Christine L
CL
D000906:Antibodies / Q000378:metabolism; D015415:Biomarkers / Q000032:analysis* / Q000378:metabolism; D015153:Blotting, Western; D002454:Cell Differentiation; D002469:Cell Separation; D053595:Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism*; D000071837:Fibrillins; D006023:Glycoproteins / Q000378:metabolism; D006801:Humans; D007553:Isotope Labeling; D013058:Mass Spectrometry; D008565:Membrane Proteins / Q000032:analysis* / Q000378:metabolism; D008840:Microfilament Proteins / Q000378:metabolism; D008856:Microscopy, Fluorescence; D032383:Myocytes, Cardiac / Q000166:cytology / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*
D000906:Antibodies; D015415:Biomarkers; C426245:EMILIN2 protein, human; D000071837:Fibrillins; D006023:Glycoproteins; D008565:Membrane Proteins; D008840:Microfilament Proteins