BMAL1-HIF2a heterodimers contribute to ccRCC [RNA-seq_Fig2]

Circadian disruption enhances cancer risk, and many tumors exhibit disordered circadian gene expression. We show rhythmic gene expression is unexpectedly robust in clear cell renal cell carcinoma (ccRCC). Furthermore, the clock gene BMAL1 is higher in ccRCC than in healthy kidneys, unlike in other tumor types. BMAL1 is closely related to ARNT and we show that BMAL1-HIF2a regulates a subset of HIF2a target genes in ccRCC cells. Depletion of BMAL1 reprograms HIF2a chromatin association and target gene expression and reduces ccRCC growth in culture and in xenografts. Analysis of pre-existing data reveals higher BMAL1 in patient-derived xenografts that are sensitive to growth suppression by HIF2a antagonists. We show that increasing BMAL1 sensitizes ccRCC-derived 786O cells to growth inhibition by PT2399. Together, these findings indicate that an alternate HIF2a heterodimer containing the circadian partner BMAL1 contributes to HIF2a activity, growth, and sensitivity to HIF2a antagonist drugs in ccRCC cells. Overall design: To measure the contributions of endogenous ARNT and BMAL1 to HIF2a-driven gene expression in ccRCC cells, we sequenced RNA prepared from 786O cells in which either ARNT or BMAL1 was depleted by shRNA. We used DESeq2 to identify transcripts that were significantly altered. We gene set enrichment analysis (GSEA) to examined the expression of 200 transcripts in the Hallmark HYPOXIA gene set.