Cancerous splice variants

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Science Signaling  17 May 2016:
Vol. 9, Issue 428, pp. ec115
DOI: 10.1126/scisignal.aag1374

Alternative splicing increases the diversity of proteins encoded by the genome. Although critical for development and to enable tissue- and cell-specific regulation and function, this process can also be pathological and contribute to disease as illustrated by a pair of papers on alternative splicing in cancer. The enzyme ketohexokinase (KHK, also known as fructokinase) metabolizes fructose, and the KHK-C splice variant is predominantly present in the liver and exhibits greater activity toward fructose than does the KHK-A variant, which is present in many tissues but has poor activity toward fructose. Li et al. found that fructose metabolism in hepatocellular carcinoma cell lines was less than that of cultured hepatocytes and that, rather than KHK-C present in the normal hepatocytes, the hepatocellular carcinoma cells had KHK-A. The splicing regulatory proteins HnRNPH1 and HnRNPH2 interacted with nucleotide sequences spanning the boundaries of spliced KHK exon, and knockdown of these RNA-binding proteins switched the isoform present in the hepatocellular carcinoma cells from KHK-A to KHK-C. Comparison of the proteins that immunoprecipitated with KHK-A and KHK-C revealed a specific interaction, which was confirmed as direct in various assays, between KHK-A and the enzyme PRPS1, which is the form of phosphoribosyl pyrophosphate synthetase that exhibits feedback inhibition and is involved in nucleotide biosynthesis. In vitro kinase assays demonstrated that KHK-A phosphorylated PRPS1 and that this phosphorylation blocked the interaction of PRPS1 with the feedback allosteric inhibitor ADP. Thus, this switch in KHK splicing in the hepatocellular carcinoma cells reduced fructose metabolism but enhanced nucleotide synthesis, which would enable proliferation. Disrupting this pathway with targeted knockdown reduced hepatocellular tumor growth of the injected cells in mice, and survival of human hepatocellular carcinoma patients negatively correlated with increased abundance of HnRNPH1 or HnRNPH2 and KHK-A and increased PRPS1 phosphorylation.

A different aspect of cancer is the ability to metastasize. MENA is an actin-regulatory protein and in vivo, but not in culture, tumor cells exhibit a switch such that the alternative splice variant MENAINV is produced. Oudin et al. found that cultured breast cancer cells engineered to express MENAINV gained the ability to migrate toward a fibronectin gradient. Intravital imaging of these cells when implanted in mice with or without a fibronectin source showed that the MENAINV-positive cells were more motile than the MENA-positive cells and that only the MENAINV-positive cells moved toward the fibronectin source. In mouse and patient tumor samples, the abundance of MENAINV correlated with reorganization of collagen in the extracellular matrix. In culture, the ability of MENAINV-positive cells to migrate on a high fibronectin gradient required the ability of fibronectin to form fibrils. Analysis of breast cancer patient data showed that the abundance of both MENA and MENAINV correlated with the amount of integrin with the α5 subunit and the amount of fibronectin and that poor outcome and relapse correlated with the abundance of MENAINV. Thus, altered alternative splicing of MENA promotes the production of an extracellular matrix–reorganizing event that enhances metastasis.

Science Signaling is interested in both the mechanisms that regulate alternative splicing and how alternative splicing influences cellular behavior. This pair of studies is an example of both sides of that research coin.

X. Li, X. Qian, L.-X. Peng, Y. Jiang, D. H. Hawke, Y. Zheng, Y. Xia, J.-H. Lee, G. Cote, H. Wang, L. Wang, C.-N. Qian, Z. Lu, A splicing switch from ketohexokinase-C to ketohexokinase-A drives hepatocellular carcinoma formation. Nat. Cell. Biol. 18, 561–571 (2016). [PubMed]

M. J. Oudin, O. Jonas, T. Kosciuk, L. C. Broye, B. C. Guido, J. Wyckoff, D. Riquelme, J. M. Lamar, S. B. Asokan, C. Whittaker, D. Ma, R. Langer, M. J. Cima, K. B. Wisinski, R. O. Hynes, D. A. Lauffenburger, P. J. Keely, J. E. Bear, F. B. Gertler, Tumor cell–driven extracellular matrix remodeling drives haptotaxis during metastatic progression. Cancer Discov. 6, 516–531 (2016). [PubMed]