Abstract 4059: Taxol inhibits HIF-1α translation by targeting HIF-1α mRNA to P-bodies following microtubule disruption

Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DC Hypoxia inducible factor-1α (HIF-1α) is a pro-angiogenic transcription factor over-expressed in over 70% of human cancers. We have previously shown that microtubule-targeting drugs (MTDs) inhibit tumor angiogenesis by downregulating HIF-1α protein and its transcriptional activity. Polysome association profiling has shown that MTDs inhibit HIF-1α by shifting its mRNA association from actively translating polysomes to the non-translating ribosomal subunits. This mechanism depends on effective drug-tubulin interaction, as no shift in HIF-1α translation is observed upon Taxol treatment in the tubulin mutant, taxol-resistant PTX10 cells, consistent with the lack of drug-induced microtubule stabilization. We next revealed a specific interaction between HIF-1α mRNA and tubulin in untreated cells through co-precipitation experiments. Moreover, visualization of HIF-1α mRNA, using HIF-specific molecular beacons, showed that HIF-1α moved bi-directionally along microtubule polymers. Interestingly, MTD treatment resulted in a significant increase in the number of Processing (P)-bodies which are cytoplasmic regions where translationally repressed mRNAs are enriched. Immunoprecipitation of the P-body component protein, Argonaute 2, revealed a 15-fold enrichment in bound HIF-1α mRNA following Taxol or other MTD treatment, suggesting that HIF-1α mRNA accumulates in P-bodies following microtubule disruption. Microtubule depolymerization by the reversible MTD, Nocodazole, also suppressed HIF-1α mRNA translation by targeting it to P-body sites; re-polymerization of microtubules, following Nocodazole washout both relieved the inhibition of HIF-1α translation and returned P-body numbers to baseline. To test whether the increase in P-body number alone was responsible for the suppression of HIF-1α translation we employed glucose starvation to increase P-body numbers without affecting cellular microtubules. Our data showed that while glucose starvation significantly increased P-body numbers, it had no effect on HIF-1α translation. Collectively, our results demonstrate that HIF-1α translation is dependent on functional microtubules, and suggest that HIF-1α mRNA may be released from P-bodies, and return to an actively translated state, once the microtubule network is restored. Our data raise the outstanding question of whether there are other mRNAs, important for cancer biology, that are similarly affected by MTD treatment. We are currently isolating translationally-active versus translationally-repressed mRNAs, by sucrose gradient centrifugation and polysome separation, from untreated or Taxol-treated MCF7 breast cancer cells, in order to identify affected genes using the Illumina Genome Analyzer IIx System. This knowledge will have important therapeutic implications given the wide use of MTDs in oncology (Supported by RO1CA114335 to PG and TL1RR024998 to MC). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4059.