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Genome-wide RNA interference analysis of renal carcinoma survival regulators identifies MCT4 as a Warburg effect metabolic target.

ABSTRACT: Clear cell renal cell carcinoma (ccRCC) is the most common pathological subtype of kidney cancer. Here, we integrated an unbiased genome-wide RNA interference screen for ccRCC survival regulators with an analysis of recurrently overexpressed genes in ccRCC to identify new therapeutic targets in this disease. One of the most potent survival regulators, the monocarboxylate transporter MCT4 (SLC16A3), impaired ccRCC viability in all eight ccRCC lines tested and was the seventh most overexpressed gene in a meta-analysis of five ccRCC expression datasets. MCT4 silencing impaired secretion of lactate generated through glycolysis and induced cell cycle arrest and apoptosis. Silencing MCT4 resulted in intracellular acidosis, and reduction in intracellular ATP production together with partial reversion of the Warburg effect in ccRCC cell lines. Intra-tumoural heterogeneity in the intensity of MCT4 protein expression was observed in primary ccRCCs. MCT4 protein expression analysis based on the highest intensity of expression in primary ccRCCs was associated with poorer relapse-free survival, whereas modal intensity correlated with Fuhrman nuclear grade. Consistent with the potential selection of subclones enriched for MCT4 expression during disease progression, MCT4 expression was greater at sites of metastatic disease. These data suggest that MCT4 may serve as a novel metabolic target to reverse the Warburg effect and limit disease progression in ccRCC.

SUBMITTER: Gerlinger M

PROVIDER: S-EPMC3504091 | biostudies-literature | 2012 Jun

REPOSITORIES: biostudies-literature

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Genome-wide RNA interference analysis of renal carcinoma survival regulators identifies MCT4 as a Warburg effect metabolic target.

Gerlinger Marco M Santos Claudio R CR Spencer-Dene Bradley B Martinez Pierre P Endesfelder David D Burrell Rebecca A RA Vetter Marcus M Jiang Ming M Saunders Rebecca E RE Kelly Gavin G Dykema Karl K Rioux-Leclercq Nathalie N Stamp Gordon G Patard Jean Jacques JJ Larkin James J Howell Michael M Swanton Charles C

The Journal of pathology 20120418 2

Clear cell renal cell carcinoma (ccRCC) is the most common pathological subtype of kidney cancer. Here, we integrated an unbiased genome-wide RNA interference screen for ccRCC survival regulators with an analysis of recurrently overexpressed genes in ccRCC to identify new therapeutic targets in this disease. One of the most potent survival regulators, the monocarboxylate transporter MCT4 (SLC16A3), impaired ccRCC viability in all eight ccRCC lines tested and was the seventh most overexpressed ge ...[more]

PMID: 22362593

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Project description:The cellular prion protein PrPC is necessary for prion replication, and its reduction greatly increases life expectancy in animal models of prion infection. Hence the factors controlling the levels of PrPC may represent therapeutic targets against human prion diseases. Here we performed an arrayed whole-transcriptome RNA interference screen to identify modulators of PrPC expression. We cultured human U251-MG glioblastoma cells in the presence of 64’752 unique siRNAs targeting 21’584 annotated human genes, and measured PrPC using a one-pot fluorescence-resonance energy transfer immunoassay in 51’128 individual microplate wells. This screen yielded 743 candidate regulators of PrPC. When downregulated, 563 of these candidates reduced and 180 enhanced PrPC expression. Recursive candidate attrition through multiple secondary screens yielded 54 novel regulators of PrPC, 9 of which were confirmed by CRISPR interference as robust regulators of PrPC biosynthesis and degradation. The phenotypes of 6 of the 9 candidates were inverted in response to transcriptional activation using CRISPRa. The RNA-binding post-transcriptional repressor Pumilio-1 was identified as a potent limiter of PrPC expression through the degradation of PRNP mRNA. Because of its hypothesis-free design, this comprehensive genetic-perturbation screen delivers an unbiased landscape of the genes regulating PrPC levels in cells, most of which were unanticipated, and some of which may be amenable to pharmacological targeting in the context of antiprion therapies. Author summary The cellular prion protein (PrPC) acts as both, the substrate for prion formation and mediator of prion toxicity during the progression of all prion diseases. Suppressing the levels of PrPC is a viable therapeutic strategy as PRNP null animals are resistant to prion disease and the knockout of PRNP is not associated with any severe phenotypes. Motivated by the scarcity of knowledge regarding the molecular regulators of PrPC biosynthesis and degradation, which might serve as valuable targets to control its expression, here, we present a cell-based genome wide RNAi screen in arrayed format. The screening effort led to the identification of 54 regulators, nine of which were confirmed by an independent CRISPR-based method. Among the final nine targets, we identified PUM1 as a regulator of PRNP mRNA by acting on the 3’UTR promoting its degradation. The newly identified factors involved in the life cycle of PrPC provided by our study may also represent themselves as therapeutic targets for the intervention of prion diseases.

Dataset Information

Genome-wide RNA interference analysis of renal carcinoma survival regulators identifies MCT4 as a Warburg effect metabolic target.

Publications

Genome-wide RNA interference analysis of renal carcinoma survival regulators identifies MCT4 as a Warburg effect metabolic target.

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