Background: Cancer cells may acquire stem cell (CSC) properties by activated TGFβ-epithelial-mesenchymal transition (EMT) axis resulting in formation of cancer stem cells. miRNAs are involved in CSC formation in solid tumors, but their role has not been investigated in renal cell carcinoma (RCC). Methods: RCC spheres were generated and propagated in serum-free defined medium (SFDM). mRNA expression was assessed by qRT-PCR. miRNA expression was screened on a qRT-PCR based panel. Tumorigenicity was assessed by subcutaneous injection of RCC sphere or parental cells into immunodeficient mice in different dilutions. TargetScan and miRPath was used for target prediction and clustering. Results: We isolated self-renewing cancer spheres from ACHN and CAKI-1 RCC cell lines in the stem cell supporting media, SFDM. Spheres were highly clonogenic and tumorigenic in xenograft tumor model and expressed high levels of stem cell-related markers and mesenchymal markers. These spheres were enriched in the mesenchymal marker CD44 and the kidney progenitor maker CD24 indicating that EMT contributed to their formation or maintenance. We compared miRNA expression between the spheres and the parental cells and identified differentially expressed miRNAs. Functional clustering of their predicted targets indicates that TGFβ signaling is a potential regulator of CSC self-renewal and is regulated by the candidate miRNAs. Further, we show that transfection of ACHN and CAKI-1 cells with the miR-17 inhibitor resulted in rapid and highly efficient formation of cancer spheres that were indistinguishable from the spheres formed in SFDM. These spheres were stable and could be propagated indefinitely. Histologic examination and immunohistochemistry of the sphere-derived xenografts confirmed the presence of clear cell RCC with large areas of sarcomatoid dedifferentiation. Finally, we prove that the TGFβ receptor II, and the co-Smad Smad4 are possible direct targets of miR-17. Conclusions: The TGFβ-EMT axis likely contributes to the self-renewing potential of RCC spheres. miRNAs are differentially expressed in RCC spheres and miR-17 inhibition transformed ccRCC cells to highly tumorigenic RCC spheres.