Abstract A13: eIF4a paralogue switching drives opposing phenotypes in cancer by specific reprogramming of gene expression
Authors: Raza Farheen , Das Madhumita , Waldron Joseph , Kopra Kari , Quesne John Le 
Published: May 2017

Aims: We aim to unravel the divergent roles of the paralogs of the ubiquitous mRNA helicase eIF4A ; these crucial proteins have emerged as major therapeutic targets in cancer.

Methods: We have used a broad range of epidemiological, tissue-based, cellular and biochemical methods. We have examined expression levels of the paralogs in a large set of lung adenocarcinomas by quantitative multiplex immunofluorescence. Cellular and biochemical effects of eIF4A paralog knockdown have been studied in cell culture using a range of growth and apoptosis assays, alongside polysome profiling, RNA-Seq methods. Reporter assays and structure-probing methods are being used to dissect how the paralogs interact with mRNA structures. Purified proteins are being used in functional assays of helicase/ATPase activity.

Introduction: Eukaryotic initiation factor 4A (eIF4A) is a ubiquitous mRNA helicase, encoded by two paralogous genes which are often considered to be functionally redundant mediators of translational initiation, although we now believe that eIF4A2 has distinct roles in the function of micro-RNAs. As eIF4A1 is known to be essential for the translation of many pro-oncogenic mRNAs, eIF4A has emerged as an attractive new cancer target.

Results: We find that eIF4A1 and eIF4A2 have extremely divergent cellular roles and clinical associations. High levels of eIF4A1 are found in proliferating/stem cell compartments of normal organs, we find that elevated eIF4A1 predicts poor outcome in lung adenocarcinoma. eIF4A2, in contrast, is expressed in terminally differentiated normal cells and has is related to good outcomes in cancer.

Therefore we have been studying the cellular effects of eIF4A1 and eIF4A2 knockdown. eIF4A1 drives cellular proliferation, while eIF4A2 is pro-aoptotic and limits cellular growth. We have identified mRNAs which are influenced by eIF4A1 and eIF4A2 at the levels of mRNA abundance and translational control. eIF4A1 is necessary for the translation of mRNAs with G/C-rich 5' UTRs and the potential for G-quadruplex formation; in contrast eIF4A2 has a generally inhibitory effect upon translation initiation, and influences a different set of mRNAs at both total and translational levels.

We find that mRNA elements with the potential for G-quadruplex formation can endow reporter mRNAs with dependence upon eIF4A1 for their translation, but that this is highly dependent upon sequence context, and also upon the size of the predicted quadruplex.

Finally, we have generated pure recombinant versions of both paralogs and their cellular binding partners, and are now starting to generate paralog-specific data on their biochemical properties.

Conclusions: eIF4A1 and eIF4A2 exhibit great divergence in their pathological, cellular and biochemical roles. eIF4A1 is associated with cancer virulence mediated by upregulation of many pro-proliferative proteins, while eIF4A2 shows the opposite tendency. The sequence and structural determinants of eIF4A-dependence are complex and highly context-dependent. eIF4A1 and eIF4A2 are biochemically distinct in functional assays. These findings are central to our understanding of the control of cellular translation, and have great implications for the therapeutic targeting of translation in cancer.

Citation Format: Farheen Raza, Madhumita Das, Joseph Waldron, Kari Kopra, John Le Quesne. eIF4a paralogue switching drives opposing phenotypes in cancer by specific reprogramming of gene expression. [abstract]. In: Proceedings of the AACR Special Conference on Translational Control of Cancer: A New Frontier in Cancer Biology and Therapy; 2016 Oct 27-30; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2017;77(6 Suppl):Abstract nr A13.