Identification of the surface-accessible, lineage-specific vascular proteome by two-dimensional peptide mapping.
The formation of blood vessels (angiogenesis) and of lymphatic vessels (lymphangiogenesis) actively contributes to cancer progression and inflammation. Thus, there has been a quest for identifying the molecular mechanisms that control lymphatic and blood vessel formation and function. Membrane and extracellular matrix proteins can serve as suitable targets for imaging and/or therapeutic targeting; however, conventional proteomic technologies often fail to identify them systematically due to insolubility in water and low abundance of membrane proteins. To circumvent this problem, we applied a gel-free proteomics methodology termed two-dimensional peptide mapping (2D-PM) to cultured blood vascular (BECs) and lymphatic (LECs) endothelial cells. 2D-PM comprises biotinylation of surface-accessible proteins, their selective enrichment, separation by HPLC, and analysis by mass spectrometry. We identified 184 proteins that were specifically or predominantly expressed by LECs and 185 proteins specifically expressed by BECs, whereas 377 additional proteins were equally detected in both cell types. For representative proteins, the differential, lineage-specific expression was confirmed by Western analyses of cultured cells and by differential immunofluorescence analyses of tissue samples. Our results identify the surface-accessible, vascular lineage-specific proteome, and they also reveal 2D-PM as a powerful technology for the large-scale screening of lineage-specific protein expression.
Roesli
Christoph
C
Institute of Pharmaceutical Sciences, Swiss Federal Institute of Technology, ETH Zurich, Wolfgang-Pauli-Str. 10, HCI H303, CH-8093 Zurich, Switzerland.
Mumprecht
Viviane
V
Neri
Dario
D
Detmar
Michael
M
D002478:Cells, Cultured; D015180:Electrophoresis, Gel, Two-Dimensional; D042783:Endothelial Cells / Q000737:chemistry*; D004729:Endothelium, Lymphatic / Q000737:chemistry*; D004730:Endothelium, Vascular / Q000737:chemistry*; D013058:Mass Spectrometry; D010449:Peptide Mapping / Q000379:methods*; D011506:Proteins / Q000032:analysis*; D020543:Proteome
D011506:Proteins; D020543:Proteome