For IRES trans-acting factors, it is all about location

Abstract

The translation of many proteins involved in transcription, cell cycle progression, apoptosis and cell survival is mediated by internal ribosome entry sites (IRESs) present within the 5′-untranslated regions (5′-UTRs) of their messenger RNA molecules (mRNAs). Several recent reports now demonstrate that the proteins controlling IRES-dependent translation initiation are regulated by their subcellular localization.

Main

Internal ribosome entry site (IRES)-mediated translation initiation is an alternative means of initiating protein synthesis that allows for the production of new protein when global, mostly cap-dependent translation initiation is attenuated (Holcik and Sonenberg, 2005). As a result, the majority of messenger RNA molecules (mRNAs) that utilize IRES for translation encode proteins that are required during conditions that decrease cap-dependent translation initiation (for instance, during mitosis, apoptosis and cellular stress). Similar to cap-dependent translation, IRES-dependent translation requires several trans-acting protein factors (collectively known as IRES trans-acting factors or ITAFs) to recruit the ribosome and initiate polypeptide synthesis (Spriggs et al., 2005). Exactly how ITAFs enable IRES-dependent translation, and how the activities of these proteins themselves are regulated, is an active field of investigation. A new report from Dobbyn et al. (2007) (this issue) and other recent findings (Lewis et al., 2007; Lin et al., 2007) suggest that the activity of ITAFs in IRES-dependent translation initiation is regulated by the subcellular distribution of these proteins.

Dobbyn et al. (2007) were studying how translation of the prosurvival protein BAG-1 is regulated during chemotoxic stress, with a particular focus on the BAG-1S isoform whose translation is mediated by an IRES. The authors found that although treatment with the chemotoxic drug vincristine represses global, cap-dependent translation initiation, the IRES-dependent translation of BAG-1S is refractory to such treatment. Subsequent experiments focused on the role of the polypyrimidine tract-binding protein (PTB) and poly(rC)-binding protein 1 (PCBP1), two known ITAFs for the BAG-1 IRES, during vincristine-resistant BAG-1S translation. Surprisingly, no changes in the abundance of these two proteins were observed following vincristine treatment, suggesting that changes in ITAF levels are not responsible for the maintenance of BAG-1 IRES-dependent translation. The authors then hypothesized that since many RNA-binding proteins (including PTB and PCBP1) are known to shuttle between the nucleus and cytoplasm, it was possible that vincristine treatment triggered the relocalization of these proteins to the cytoplasm, where they can facilitate the IRES-dependent translation of BAG-1S. Indeed, the authors found that both PTB and PCBP1 accumulate in the cytoplasm following vincristine treatment, suggesting that regulated subcellular relocalization of ITAFs is an important determinant in IRES-dependent translation.

Other recent reports have also emphasized the importance of subcellular relocalization for ITAF function. Lin et al. (2007) identified RNA-binding motif protein 4 (RBM4), which has a regulatory function in alternative splicing of pre-mRNA, as a novel ITAF that can assist in the recruitment of the RNA helicase eIF4A to IRES-containing mRNAs to enable translation initiation. Interestingly, the subcellular relocalization of RBM4 from the nucleus to the cytoplasm is critical for RBM4 ITAF activity. The authors also found that RBM4 subcellular localization is controlled by the MKK3/6-p38 signaling pathway, which is responsive to mitogens and extracellular stresses (Ono and Han, 2000). It might then be expected that RBM4 cytoplasmic accumulation results in the upregulation of IRES that control the translation of proteins involved in proliferation and the stress response. Indeed, cytoplasmic accumulation of RBM4 was found to enhance the recruitment of eIF4A to the c-myc IRES and the Bcl-2 IRES, which mediate the translation of a transcription factor and an antiapoptotic protein, respectively.

Similarly, Lewis et al. (2007) found that subcellular relocalization of an ITAF plays a critical role in IRES-dependent translation. These investigations showed that cytoplasmic accumulation of hnRNP A1 represses the IRES-dependent translation of the X-linked inhibitor of apoptosis (XIAP), indicating that the subcellular relocalization of ITAFs can have a negative effect on IRES-dependent translation. Moreover, the cytoplasmic relocalization of an ITAF can have opposing effects on IRES-dependent translation depending on the IRES targeted by the ITAF. This was shown by the observation that cytoplasmic accumulation of hnRNP A1 enhances the IRES-dependent translation of fibroblast growth factor 2 (FGF-2) but represses the IRES-dependent translation of XIAP (Lewis et al., 2007).

Many ITAFs participate in pre-mRNA processing events such as alternative splicing when present in the nucleus (Mayeda and Krainer, 1992; Singh et al., 1995), yet are involved in IRES-dependent translation when present in the cytoplasm. It would therefore appear that the diverse roles of ITAFs in mRNA metabolism are controlled at least partially by the compartmentalization of these proteins. The realization that the shuttling of ITAFs between the nucleus and the cytoplasm affects their activity suggests two mechanisms by which ITAF subcellular distribution may control IRES-dependent translation (Figure 1). In one case, the nuclear localized ITAF may associate with its target IRES-containing mRNA in this compartment, resulting in the majority of the transcript being sequestered in the nucleus and therefore separated from the translational machinery. Following the appropriate signal, the ITAF accumulates in the cytoplasm, carrying with it the IRES-containing mRNA, where the ITAF may actively participate in the recruitment of the ribosome. Alternatively, the ITAF may be primarily located in the nucleus to keep it separate from its target IRES and the translational machinery until such time that translation of the IRES-containing mRNA is warranted and a signal causes the ITAF to accumulate in the cytoplasm. Experiments that address the subcellular distribution of IRES-containing mRNAs following the knockdown of a shuttling ITAF should help to clarify the mechanism(s) involved.

Figure 1

Two proposed models for the regulation of internal ribosome entry site (IRES) trans-acting factor (ITAF) function by subcellular localization. (a) ITAFs are normally localized in the nucleus, where they associate with their target IRES-containing messenger RNA molecule (mRNA; IRES mRNA), resulting in the sequestration of the majority of the transcripts in the nucleus, and consequently poor translation. Following the appropriate signal (such as treatment with chemotoxic drugs) the ITAF–RNA complexes accumulate in the cytoplasm, where the ITAFs actively participate in the recruitment of the ribosome and enhanced IRES-mediated translation. (b) Alternatively, ITAFs are located in the nucleus to keep them separate from the target IRES mRNA and the translational machinery. Following the appropriate signal, the ITAFs accumulate in the cytoplasm where they associate with their target IRES mRNAs and enable efficient IRES-dependent translation. In both models, the conditions that stimulate IRES-mediated translation attenuate global, cap-dependent translation. (ITAFs are depicted as red circles, the m7G cap as a small black circle, IRES as white rectangles and non-IRES mRNAs are green.)

Although it may be intuitive that proteins that are involved in translation initiation must be localized in the cytoplasm to function, the ability to separate an ITAF from its target IRES-containing mRNA and/or the ribosomal machinery may be a simple, yet important, regulatory mechanism for IRES-dependent translation. Moreover, many mRNAs whose translation is mediated by IRES encode proteins involved in critical cellular processes, and it is therefore important that the production of these proteins be tightly regulated. In fact, it is entirely possible that the inappropriate subcellular distribution of ITAFs contributes to the dysregulation of IRES-dependent translation during disease and pathophysiological conditions. This suggestion is highlighted by the fact that Dobbyn et al. (2007) find that treatment of cells with the chemotoxic drug vincristine in fact increases the relative abundance of the antiapoptotic factor BAG-1S due to reinforced IRES-mediated translation caused by the vincristine-induced subcellular relocalization of PTB and PCBP1. Thus, cells that should succumb to vincristine treatment are instead resistant to the drug due to enhanced IRES-dependent translation of an antiapoptotic factor. This observation may therefore account for the chemoresistance of cancer cells in response to some chemotoxic drugs, as many antiapoptotic proteins depend on IRES for their translation (Baird et al., 2006) and the activity of these IRES may be enhanced by ITAFs that relocalize to the cytoplasm following treatment with such therapeutic compounds.

The fact that several ITAFs shuttle between the nucleus and the cytoplasm, and that in many cases this distribution is likely controlled by extracellular cues, indicates that the subcellular distribution of ITAFs may be a widely utilized mechanism to control IRES-dependent translation. In addition to mammalian cells, many viruses also make efficient use of IRES-dependent translation and it may therefore be interesting to determine if viral infection activates signaling pathways that induce the subcellular redistribution of ITAFs, which in turn enhance viral IRES activity. Future investigations that focus on the elucidation of signaling pathways that control the subcellular distribution of ITAFs should provide a clearer picture of how IRES-dependent translation is regulated.