Loss of VHL and hypoxia provokes PAX2 up-regulation in clear cell renal cell carcinoma.
OBJECTIVE
The paired box gene 2, PAX2, encodes for a transcription factor that is up-regulated during nephrogenesis and becomes silenced in mature epithelium of the glomeruli, the proximal, and distal tubules. Reactivation of PAX2 has been frequently observed in clear cell renal cell carcinoma (ccRCC), a tumor type characterized by loss of von Hippel-Lindau (VHL) tumor suppressor function. The regulation of PAX2 expression in ccRCC is unknown.


METHODS
We applied reporter gene assays to investigate PAX2 promoter regulation. Furthermore, PAX2 expression was determined in ccRCC cell lines under normoxic and hypoxic condition in a VHL wild-type and mutated background. PAX2 expression was also assessed in 831 human ccRCC and correlated with hypoxia-inducible factor alpha (HIFalpha) and clinical parameters.


RESULTS
Here, we show that both loss of VHL protein (pVHL) function and hypoxia leads to strong PAX2 reexpression. Using luciferase reporter gene assays, no induction was obtained in spite of six hypoxia response element motifs identified in the promoter of PAX2. Comprehensive immunohistochemical analyses showed significant correlations between PAX2, HIF1alpha, and HIF2alpha-target CCND1 expression patterns in ccRCC patients. Notably, PAX2 expression was highly associated with early-stage, well-differentiated ccRCC and, consequently, better clinical outcome (P < 0.0001 each). Additional analyses indicated that PAX2 repressor WT1 and cancer-linked hypomethylation are not important for transcriptional regulation of PAX2 in ccRCC.


CONCLUSIONS
We conclude that in ccRCC, PAX2 reactivation is driven by HIF-dependent mechanisms following pVHL loss.
Luu
Van-Duc
VD
Department of Pathology, Institute of Surgical Pathology, University Hospital Zurich, Zurich, Switzerland.
Boysen
Gunther
G
Struckmann
Kirsten
K
Casagrande
Silvia
S
von Teichman
Adriana
A
Wild
Peter J
PJ
Sulser
Tullio
T
Schraml
Peter
P
Moch
Holger
H
D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics / Q000378:metabolism; D015153:Blotting, Western; D002292:Carcinoma, Renal Cell / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D015687:Cell Hypoxia; D002460:Cell Line; D045744:Cell Line, Tumor; D018899:CpG Islands / Q000235:genetics; D019175:DNA Methylation; D006801:Humans; D007150:Immunohistochemistry; D007680:Kidney Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D009367:Neoplasm Staging; D051762:PAX2 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016019:Survival Analysis; D015854:Up-Regulation; D051794:Von Hippel-Lindau Tumor Suppressor Protein / Q000235:genetics / Q000378:metabolism*; D000099162:Endothelial PAS Domain-Containing Protein 1
D051792:Basic Helix-Loop-Helix Proteins; D008156:Luciferases; D051762:PAX2 Transcription Factor; D011993:Recombinant Fusion Proteins; D051794:Von Hippel-Lindau Tumor Suppressor Protein; D000099162:Endothelial PAS Domain-Containing Protein 1; C497318:PAX2 protein, human