The Axl-Regulating Tumor Suppressor miR-34a Is Increased in ccRCC but Does Not Correlate with Axl mRNA or Axl Protein Levels.
BACKGROUND
High expression of the receptor tyrosine kinase Axl is associated with poor prognosis in patients with Renal Cell Carcinoma (RCC), the most common malignancy of the kidney. The miR-34a has been shown to directly regulate Axl in cancer cells. The miR-34a is a mediator of p53-dependent tumor suppression, and low expression of miR-34a has been associated with worse prognosis in several cancers. Our aim was to elucidate whether miR-34a or the other members of the miR-34 family (miR-34b/c) regulate Axl in RCC.


RESULTS
Using western blot, flow cytometry, and RT-qPCR, we showed that Axl mRNA and protein are downregulated in 786-O cells by miR-34a and miR-34c but not by miR-34b. A luciferase reporter assay demonstrated direct interaction between the Axl 3' UTR and miR-34a and miR-34c. The levels of miR-34a/b/c were measured in tumor tissue in a cohort of 198 RCC patients, and the levels of miR-34a were found to be upregulated in clear cell RCC (ccRCC) tumors, but not associated with patient outcome. Neither of the miR-34 family members correlated with Axl mRNA, soluble Axl protein in serum, nor with immunohistochemistry of Axl in tumor tissue. In addition, we measured mRNA levels of a known miR-34a target, HNF4A, and found the HNF4A levels to be decreased in ccRCC tumors, but unexpectedly correlated positively rather than negatively with miR-34a.


CONCLUSIONS
Although miR-34a and miR-34c can regulate Axl expression in vitro, our data indicates that the miR-34 family members are not the primary regulators of Axl expression in RCC.
Lund University, Department of Translational Medicine, Section of Clinical Chemistry, University Hospital Malmö, Malmö, Sweden.
Helena K
HK
Fritz
Lund University, Department of Translational Medicine, Section of Clinical Chemistry, University Hospital Malmö, Malmö, Sweden.
Anna
A
Gustafsson
Umeå University, Departments of Surgical and Perioperative Sciences, Urology and Andrology, Umeå, Sweden.
Börje
B
Ljungberg
Lund University, Department of Laboratory Medicine, Division of Translational Cancer Research, Medicon Village, Lund, Sweden.
Yvonne
Y
Ceder
Lund University, Department of Laboratory Medicine, Division of Translational Cancer Research, Medicon Village, Lund, Sweden.
Håkan
H
Axelson
Lund University, Department of Translational Medicine, Section of Clinical Chemistry, University Hospital Malmö, Malmö, Sweden.
Björn
B
Dahlbäck
D002292:Carcinoma, Renal Cell / Q000235:genetics* / Q000401:mortality; D045744:Cell Line, Tumor; D015972:Gene Expression Regulation, Neoplastic; D016147:Genes, Tumor Suppressor; D057809:HEK293 Cells; D051557:Hepatocyte Nuclear Factor 4 / Q000235:genetics*; D006801:Humans; D035683:MicroRNAs / Q000235:genetics*; D011379:Prognosis; D011518:Proto-Oncogene Proteins / Q000235:genetics* / Q000378:metabolism; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics* / Q000378:metabolism; D000094963:Axl Receptor Tyrosine Kinase
C496592:HNF4A protein, human; D051557:Hepatocyte Nuclear Factor 4; C522425:MIRN34 microRNA, human; D035683:MicroRNAs; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D020794:Receptor Protein-Tyrosine Kinases; D000094963:Axl Receptor Tyrosine Kinase; C000722289:AXL protein, human