No abstract

Initiation of translation in eukaryotic cells can occur by two distinct mechanisms, cap-dependent scanning and internal ribosome entry. The latter mechanism requires the formation of a complex RNA structural element termed an internal ribosome entry segment (IRES). IRESs are located in the 5 0 untranslated region of the message, and in the presence of trans-acting factors allow the ribosome to be recruited to a site that is a considerable distance from the cap structure. Many cellular mRNAs have now been shown to contain IRESs and it is likely that up to 10% of all mRNAs have the capability to initiate translation by this mechanism. The majority of IRESs that have been identified thus far are found in mRNAs whose protein products are associated with the control of cell growth and cell death, including many growth factors, proto-oncogenes and proteins required for apoptosis. In this review, we discuss the cellular situations when IRESs are required, the trans-acting factors that are necessary for IRES function and deregulation of IRES-mediated translation in tumorigenesis.

There is now an overwhelming body of evidence to suggest that internal ribosome entry is required to maintain the expression of specific proteins during patho‐physiological situations when cap‐dependent translation is compromised, for example, following heat shock or during mitosis, hypoxia, differentiation and apoptosis. Translational profiling has been used by several groups to assess the extent to which alternative mechanisms of translation initiation selectively recruit mRNAs to polysomes during cell stress. The data from these studies have shown that under each condition 3–5% of coding mRNAs remain associated with the polysomes. Importantly, the genes identified in each of these studies do not show a significant amount of overlap, suggesting that 10–15% of all mRNAs have the capability for their initiation to occur via alternative mechanism(s).

Recent studies have shown that during apoptosis protein synthesis is inhibited and that this is in part due to the proteolytic cleavage of eukaryotic initiation factor 4G (eIF4G). Initiation of translation can occur either by a cap-dependent mechanism or by internal ribosome entry. The latter mechanism is dependent on a complex structural element located in the 5 untranslated region of the mRNA which is termed an internal ribosome entry segment (IRES). In general, IRES-mediated translation does not require eIF4E or full-length eIF4G. In order to investigate whether cap-dependent and cap-independent translation are reduced during apoptosis, we examined the expression of c-Myc during this process, since we have shown previously that the 5 untranslated region of the c-myc proto-oncogene contains an IRES. c-Myc expression was determined in HeLa cells during apoptosis induced by tumor necrosis factor-related apoptosis-inducing ligand. We have demonstrated that the c-Myc protein is still expressed when more than 90% of the cells are apoptotic. The presence of the protein in apoptotic cells does not result from either an increase in protein stability or an increase in expression of c-myc mRNA. Furthermore, we show that during apoptosis initiation of c-myc translation occurs by internal ribosome entry. We have investigated the signaling pathways that are involved in this response, and cotransfection with plasmids which harbor either wild-type or constitutively active MKK6, a specific immediate upstream activator of p38 mitogen-activated protein kinase (MAPK), increases IRES-mediated translation. In addition, the c-myc IRES is inhibited by SB203580, a specific inhibitor of p38 MAPK. Our data, therefore, strongly suggest that the initiation of translation via the c-myc IRES during apoptosis is mediated by the p38 MAPK pathway.

The 5' UTR of c -myc mRNA contains an internal ribo-some entry segment (IRES) and consequently, c -myc mRNAs can be translated by the alternative mechanism of internal ribosome entry. However, there is also some evidence suggesting that c -myc mRNA translation can occur via the conventional cap-dependent scanning mechanism. Using both bicistronic and monocistronic mRNAs containing the c- myc 5' UTR, we demonstrate that both mechanisms can contribute to c- myc protein synthesis. A wide range of cell types are capable of initiating translation of c- myc by internal ribosome entry, albeit with different efficiencies. Moreover, our data suggest that the spectrum of efficiencies observed in these cell types is likely to be due to variation in the cellular concentration of non-canonical translation factors. Interestingly, the c -myc IRES is 7-fold more active than the human rhinovirus 2 (HRV2) IRES and 5-fold more active than the encephalomyocarditis virus (EMCV) IRES. However, the protein requirements for the c -myc IRES must differ significantly from these viral IRESs, since an unidentified nuclear event appears to be a pre-requisite for efficient c -myc IRES-driven initiation.