The most common genetic aberration associated with inherited or sporadic renal cell carcinoma (RCC) is the loss or nonfunctional mutations in the von Hippel-Lindau (VHL) tumor suppressor gene. The mutations lead to the stabilization of hypoxia inducible factor (HIF-1α and 2α), thereby promoting the expression of HIF target genes. HIF target genes are reported to induce tumor phenotypes by altering cellular metabolism and upregulating the expression of protumorigenic/angiogenic factors as well. We report here tissue microarray data and histology-based analyses demonstrating that the gene PRDM16 (PRD1-BF1-RIZ1 homologous domain containing 16) is silenced in many RCC tumors regardless of their VHL status. Bioinformatics analysis demonstrates a role for promoter methylation in silencing PRDM16. We investigated the role of PRDM16 in the context of VHL loss in kidney cancer. PRDM16 is known to regulate brown fat mitochondrial metabolism by inducing expressions of peroxisome proliferator-activated receptor-gamma coactivator-1alpha (PPARGC1A) and estrogen-related receptor-gamma (ESRRG). Altered cellular metabolism, in particular, decreased mitochondrial respiration is a hallmark of RCC, which is in attributed to VHL alterations. Analysis of TCGA data demonstrates that PRDM16 expression strongly correlates with PPARGC1A expression in RCC. Ectopic expression of PRDM16 in RCC cells can induce PPARGC-1α and ESRRG mRNA expression but fails to increase mitochondrial respiration. However, ectopic expression of PRDM16 in RCC cell lines can suppress both in vitro and in vivo tumor phenotypes. These data suggest that the tumor-suppressive activity of PRDM16 may be independent from its effects on mitochondrial metabolism. RNA-seq based analysis of a VHL mutated RCC line re-expressing PRDM16 demonstrates that a transcription suppressive role of PRDM16 predominates in RCC. PRDM16 suppresses genes involved in cell migration and proliferation. In particular, PRDM16 downregulates the expression of semaphorin 5B (SEMA5B). q-PCR analysis validates high-throughput data in a panel of RCC and embryonic kidney cell lines. SEMA5B knock down significantly decreases RCC proliferation supporting the protumorigenic function of SEMA5B in RCC. Given the central role of VHL loss in renal tumor initiation, we demonstrate that the VHL/HIF axis regulates the expression of SEMA5B in RCC. Collectively, these data suggest that transcription suppressive activity of PRDM16 and VHL controls expression of the protumorigenic factor SEMA5B and that epigenetic silencing of PRDM16 thus amplifies this consequence of VHL loss.

Citation Format: Anirban Kundu, Eun-Young Kho, Sandeep B. Shelar, Hyeyoung Nam, Garret Brinkley, Shimoga Darshan, Yawen Tang, Richard Kirkman, David K. Crossman, Sooryanarayana Varambally, Glenn C. Rowe, Shi Wei, Phillip Buckhaults, Sunil Sudarshan. Functional implications of PRDM16 loss in kidney cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4483.