von Hippel-Lindau tumor suppressor gene-dependent mRNA stabilization of the survival factor parathyroid hormone-related protein in human renal cell carcinoma by the RNA-binding protein HuR.
We have shown that parathyroid hormone-related protein (PTHrP) is a survival factor for human renal cell carcinoma (RCC) and that its expression is negatively regulated by the von Hippel-Lindau (VHL) tumor suppressor gene at the level of messenger RNA (mRNA) stability, as observed for tumor growth factors (TGFs). Our goals were to analyze the alternative splicing of PTHrP mRNA in human RCC and from these results to identify VHL/hypoxia-induced factor (HIF) system-regulated mRNA-binding proteins involved in PTHrP mRNA stability. We used: (i) a panel of human RCC cells expressing or not VHL; (ii) VHL-deficient 786-0 cells transfected with active or inactive VHL and (iii) human RCC samples and corresponding normal tissues. By quantitative real-time reverse transcription-polymerase chain reaction analysis, the 141 PTHrP mRNA isoform was found to be predominant in all cells and tumors (80%). In cells transfected with VHL, the expressions of all isoforms were decreased by 50%. Eight proteins with molecular weights ranging from 20 to 75 kDa were found to bind to biotinylated transcripts spanning the 141 PTHrP mRNA AU-rich 3'-untranslated region whose abundancy was dependent on VHL expression. The protein having an apparent molecular weight of 30 kDa was identified by western blot as HuR, a RNA-binding protein with stabilizing functions on various mRNA coding for proteins important in malignant transformation including vascular endothelial growth factor and TGF-beta. PTHrP expression studies confirmed the involvement of HuR in PTHrP upregulation in this disease. Common mRNA-binding proteins regulated by the VHL/HIF system may constitute new therapeutic opportunities against human RCC that remains refractory to therapies.
Danilin
Sabrina
S
Institut National de la Santé et de la Recherche Médicale U727, Section of Renal Pharmacology and Physiopathology, School of Medicine, University Louis Pasteur, Strasbourg, France.
Sourbier
Carole
C
Thomas
Lionel
L
Rothhut
Sylvie
S
Lindner
Véronique
V
Helwig
Jean-Jacques
JJ
Jacqmin
Didier
D
Lang
Hervé
H
Massfelder
Thierry
T
D017398:Alternative Splicing; D000954:Antigens, Surface / Q000378:metabolism*; D002292:Carcinoma, Renal Cell / Q000378:metabolism*; D045744:Cell Line, Tumor; D051959:ELAV Proteins; D000067780:ELAV-Like Protein 1; D006801:Humans; D007680:Kidney Neoplasms / Q000378:metabolism*; D044162:Parathyroid Hormone-Related Protein / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D020871:RNA Stability / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D051794:Von Hippel-Lindau Tumor Suppressor Protein / Q000502:physiology*
D000954:Antigens, Surface; D051959:ELAV Proteins; D000067780:ELAV-Like Protein 1; C098770:ELAVL1 protein, human; D044162:Parathyroid Hormone-Related Protein; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D051794:Von Hippel-Lindau Tumor Suppressor Protein