Identification and functional characterization of pVHL-dependent cell surface proteins in renal cell carcinoma.
The identification of cell surface accessible biomarkers enabling diagnosis, disease monitoring, and treatment of renal cell carcinoma (RCC) is as challenging as the biology and progression of RCC is unpredictable. A hallmark of most RCC is the loss-of-function of the von Hippel-Lindau (pVHL) protein by mutation of its gene (VHL). Using the cell surface capturing (CSC) technology, we screened and identified cell surface N-glycoproteins in pVHL-negative and positive 786-O cells. One hundred six cell surface N-glycoproteins were identified. Stable isotope labeling with amino acids in cell culture-based quantification of the CSC screen revealed 23 N-glycoproteins whose abundance seemed to change in a pVHL-dependent manner. Targeted validation experiments using transcriptional profiling of primary RCC samples revealed that nine glycoproteins, including CD10 and AXL, could be directly linked to pVHL-mediated transcriptional regulation. Subsequent human tumor tissue analysis of these cell surface candidate markers showed a correlation between epithelial AXL expression and aggressive tumor phenotype, indicating that pVHL-dependent regulation of glycoproteins may influence the biologic behavior of RCC. Functional characterization of the metalloprotease CD10 in cell invasion assays demonstrated a diminished penetrating behavior of pVHL-negative 786-O cells on treatment with the CD10-specific inhibitor thiorphan. Our proteomic surfaceome screening approach in combination with transcriptional profiling and functional validation suggests pVHL-dependent cell surface glycoproteins as potential diagnostic markers for therapeutic targeting and RCC patient monitoring.
Institute of Surgical Pathology, University Hospital Zurich, Zurich, Switzerland.
Gunther
G
Boysen
Damaris
D
Bausch-Fluck
Claudio R
CR
Thoma
Anna M
AM
Nowicka
Daniel P
DP
Stiehl
Igor
I
Cima
Van-Duc
VD
Luu
Adriana
A
von Teichman
Thomas
T
Hermanns
Tullio
T
Sulser
Barbara
B
Ingold-Heppner
Niklaus
N
Fankhauser
Roland H
RH
Wenger
Wilhelm
W
Krek
Peter
P
Schraml
Bernd
B
Wollscheid
Holger
H
Moch
D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics / Q000378:metabolism; D002292:Carcinoma, Renal Cell / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D015972:Gene Expression Regulation, Neoplastic; D006023:Glycoproteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007680:Kidney Neoplasms / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D015260:Neprilysin / Q000097:blood / Q000235:genetics / Q000378:metabolism; D040901:Proteomics; D015203:Reproducibility of Results; D051794:Von Hippel-Lindau Tumor Suppressor Protein / Q000378:metabolism*
D051792:Basic Helix-Loop-Helix Proteins; D006023:Glycoproteins; D008565:Membrane Proteins; D015260:Neprilysin; D051794:Von Hippel-Lindau Tumor Suppressor Protein; C497446:VHL protein, human